Oxazolidinone liposome composition

Abstract

One aspect of the present disclosure relates to various liposomal compositions of oxazolidinone compounds, as well as related methods of making and using the oxazolidinone liposomal compositions. In some embodiments, the liposomal compositions have improved storage stability with respect to degradation of components.

Description

[Technical Field]

[0001] [Related Applications] This patent application claims the benefit of and priority to U.S. Provisional Patent Application No. 63 / 292,899, filed December 22, 2021, which is incorporated herein by reference in its entirety.

[0002] The present disclosure relates to liposomal compositions containing oxazolidinone compounds, methods for making the same, and the use of aminoalkyl oxazolidinone compounds in the treatment of Mycobacterium tuberculosis and other gram-positive bacterial infections. [Background technology]

[0003] Liposome composition is useful for delivering therapeutic compounds.Liposome composition can comprise liposome that encapsulates therapeutic compounds in the vesicle formed by the membrane formed by lipid.Liposome is usually characterized by having an internal space that is separated from the external medium by one or more bilayer membranes that form a microscopic sac or vesicle.

[0004] However, liposomes encapsulating therapeutic compounds may degrade during storage and before therapeutic administration. For example, during storage of liposome compositions containing therapeutic compounds, oxidative degradation of liposome components and changes in liposome particle size or polydispersity index (PDI) may occur. There remains a need for liposome compositions containing therapeutic agents that have improved storage stability and demonstrate reduced liposome degradation during storage. Summary of the Invention

[0005] Provided herein are liposome compositions and methods for treating methicillin-resistant Staphylococcus aureus (MRSA) bacterial infections.

[0006] One aspect of the present disclosure is a compound of formula (I): [ka] wherein R1 is a tetrazole ring substituted at the 2' position with an aminoalkyl; and R2 is amino or acetamido. or a pharmaceutically acceptable salt thereof, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is encapsulated in liposomes in an aqueous medium having a pH greater than 6.7, and the liposomes contain phosphatidylcholine, cholesterol, and a PEG polymer-conjugated lipid, and the cholesterol content is 50 to 65 mol % relative to the total of cholesterol and non-PEGylated phospholipid in the liposomes.

[0007] In some embodiments, R2 is acetamide (NHCOCH3). In some embodiments, R1 is [ka] is selected from the group consisting of:

[0008] In some embodiments, the PEG polymer-conjugated lipid is present in an amount of 5 mol% relative to the phosphatidylcholine. In some embodiments, the sulfate salt of the compound of Formula (I) is encapsulated in liposomes comprising phosphatidylcholine, cholesterol, and the PEG polymer-conjugated lipid in a molar ratio of 45:55:2.25. In some embodiments, the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soybean phosphatidylcholine (HSPC). In some embodiments, the PEG polymer-conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE). In some embodiments, the liposome composition further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA, and the chelating agent is present at a concentration of 0.1 to 1 mM.

[0009] In some embodiments, the compound of Formula (I) is a compound selected from AKG-38, AKG-39, and AKG-40, or a pharmaceutically acceptable salt thereof. [Table 1]

[0010] In some embodiments, the compound of formula (I) is the sulfate salt of AKG-38.

[0011] In some embodiments, the liposome composition has a pH greater than 7 and equal to or less than 8. In some embodiments, the liposome composition has a pH of 7.3 to 7.7. In some embodiments, the liposome composition has a pH of 7.5.

[0012] In some embodiments, the compound of formula (I) is a sulfate salt of AKG-38; [ka] The compound is encapsulated in liposomes formed from hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 in an aqueous medium at a pH of 7.3 to 7.7. In some embodiments, the liposome composition further comprises a chelating agent, which is deferoxamine (DFO), at a concentration of 0.1 to 1 mM. In some embodiments, the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 430 to 680 g / mol. In some embodiments, the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 600 g / mol. In some embodiments, the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm; the liposome composition has a polydispersity index of less than 0.15. In some embodiments, the liposome composition has a ratio of encapsulated AKG-38 to total AKG-38 of at least 90%. In some embodiments, the aqueous medium further comprises sodium chloride. In some embodiments, the aqueous medium has an osmolality of 270-330 mOsmol / kg; the sodium chloride is at a concentration of 130-150 mM; and the chelating agent is at a concentration of 0.5 mM. In some embodiments, the aqueous medium comprises ammonium ions at a concentration of 20-60 mM, and the sodium chloride is at a concentration of 50-80 mM. In some embodiments, the liposome composition further comprises HEPES or a phosphate buffer.

[0013] One aspect of the present disclosure relates to an AKG-38 liposomal composition having a pH of at least 7.0 and not more than 8.0, the liposomal composition comprising the lipids HSPC, cholesterol, and PEG(2000)-DSPE, and AKG-38 in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1. [ka] and the liposome composition further comprises a pharmaceutically acceptable salt of: (a) the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm, or the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 100 to 130 nm; (b) the liposome composition has a polydispersity index of less than 0.15, or the liposome composition has a polydispersity index of less than 0.10; (c) the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 430 to 480 g / mol, or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 500 to 650 g / mol; or the AKG- (d) the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 430 to 650 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 450 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 450 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 600 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 600 g / mol; (d) the total concentration of AKG-38 in the liposome composition is 12 to 25 mg / mL, or the total concentration of AKG-38 in the liposome composition is 13.5 to 16 mg / mL.(e) the ratio of encapsulated AKG-38 to total AKG-38 in the liposome composition is at least 90%, or the ratio of encapsulated AKG-38 to total AKG-38 in the liposome composition is at least 95%, or the ratio of encapsulated AKG-38 to total AKG-38 in the liposome composition is at least 5 mg / mL, or the total concentration of AKG-38 in the composition is 15 mg / mL, or the total concentration of AKG-38 in the liposome composition is 20 mg / mL (200 mg in a 10 mL vial); (f) the liposome composition comprises an aqueous medium containing sodium chloride and optionally ammonium ions; (g) the aqueous medium has an osmolality of 270 to 330 mOsmol / kg, or the aqueous medium has an osmolality of 270 to 310 mOsmol / kg; (h) the aqueous medium contains ammonium ions at a concentration of 20 to 60 mM, or the aqueous medium contains ammonium ions at a concentration of 50 to 80 mM. (i) the aqueous medium contains sodium chloride at a concentration of 130 to 150 mM; (j) the aqueous medium further contains a buffer solution, which buffers the liposome composition at a pH of 7.3 to 7.7, or a pH of 7.5; (k) the aqueous medium further contains HEPES or a phosphate buffer, or the aqueous medium contains ammonium ions at a concentration of 5 to 50 mM or the aqueous medium further comprises HEPES or phosphate buffer at a concentration of 20 mM; (l) the aqueous medium further comprises a chelating agent, or the aqueous medium further comprises a chelating agent at a concentration of 0.1 to 1 mM, or the aqueous medium further comprises a chelating agent at a concentration of 0.5 mM, or the aqueous medium further comprises deferoxamine (DFO) or EDTA, or the aqueous medium further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM, or the aqueous medium 0.(m) the liposome composition is storage stable; or (n) the AKG-38 is encapsulated in the liposomes as the sulfate salt of AKG-38.

[0014] One aspect of the present disclosure is a method for the preparation of a chelating agent selected from the group consisting of deferoxamine (desferrioxamine, desferral), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), comprising administering to the patient a chelating agent (5R )-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one, or a pharmaceutically acceptable salt thereof, and encapsulated in liposomes comprising hydrogenated soy phosphatidylcholine (HSPC), cholesterol, and PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine, PEG-DSPE (PEG(2000)-DSPE).

[0015] In some embodiments, the isotonic AKG-38 liposome dispersion has a pH greater than 6.7 and less than or equal to 8.0. In some embodiments, the isotonic AKG-38 liposome dispersion has a pH of 7.5. In some embodiments, the liposomes are formed from hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.2. In some embodiments, the chelating agent is deferoxamine. In some embodiments, the liposome dispersion comprises lipid vesicles formed from a composition comprising phosphatidylcholine, 55 mol% cholesterol, and 5 mol% PEG-DSG or 5 mol% PEG-DSPE.

[0016] One aspect of the present disclosure relates to a method of treating a methicillin-resistant Staphylococcus aureus (MRSA) bacterial infection, the method comprising administering to a subject in need thereof a therapeutically effective amount of a liposome composition.

[0017] One aspect of the present disclosure relates to a method for making a liposome composition, the method comprising the steps of: (a) dissolving one or more phospholipids, cholesterol, and a PEG-lipid derivative in ethanol to obtain a lipid solution; (b) combining the lipid solution of step (a) with a sequestering agent solution to obtain a homogenous lipid suspension having a desired phospholipid concentration; (c) extruding the lipid suspension of step (b) through a membrane having a defined pore size, such as a polycarbonate track-etched (PCTE) membrane having a nominal pore size of 50-200 nm; and (d) separating the lipids in the extruded lipid suspension. (e) contacting the liposomes with a compound of formula (I) in an aqueous medium to effect encapsulation of the compound in the liposomes; (f) optionally removing any unencapsulated compound; and (g) providing the liposomes in a physiologically acceptable medium suitable for parenteral use, wherein the sequestering agent solution of step (b) comprises an aqueous ammonium sulfate solution at a concentration greater than 0.25 M, and the physiologically acceptable medium of step (g) comprises a chelating agent.

[0018] In some embodiments, the capture agent solution in step (b) comprises ammonium sulfate at a concentration of 0.5 M. In some embodiments, the chelating agent is deferoxamine. In some embodiments, the extruded liposomes in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm.

[0019] Provided herein are liposome compositions and methods for treating mycobacterial infections.

[0020] In some embodiments, the liposome composition comprises a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein R2 is an amine (NH2). In some embodiments, R1 is [ka] is selected from the group consisting of:

[0021] In some embodiments, the compound of Formula (I) or a pharmaceutically acceptable salt thereof is encapsulated in liposomes in an aqueous medium having a pH greater than 6.7; the liposomes comprise phosphatidylcholine, cholesterol, and a PEG polymer-conjugated lipid, with the cholesterol content being 50-65 mol% relative to the total cholesterol and non-PEGylated phospholipid content in the liposome. In some embodiments, the PEG polymer-conjugated lipid is present in an amount of 5 mol% relative to the phosphatidylcholine. In some embodiments, the sulfate salt of the compound of Formula (I) is encapsulated in liposomes comprising phosphatidylcholine, cholesterol, and a PEG polymer-conjugated lipid in a molar ratio of 45:55:2.25. In some embodiments, the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soybean phosphatidylcholine (HSPC). In some embodiments, the PEG polymer-conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE). In some embodiments, the liposome composition further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA, wherein the chelating agent is at a concentration of 0.1 to 1 mM.

[0022] In some embodiments, the compound of Formula (I) is a compound selected from AKG-28, AKG-29, AKG-30, AKG-31, AKG-38, and AKG-39, or a pharmaceutically acceptable salt thereof. [Table 2]

[0023] In some embodiments, the compound of formula (I) is AKG-28 [ka] The compound is a sulfate salt of PEG-DSPE, and the compound is encapsulated in liposomes formed from hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 in an aqueous medium with a pH of 7.3-7.7.

[0024] One aspect of the present disclosure relates to an AKG-28 liposomal composition comprising liposomes, the liposomes being composed of the lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1, and AKG-28 encapsulated in the liposomes. [ka] and the liposome composition further comprises a pharmaceutically acceptable salt thereof, wherein (a) the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm, or the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 100 to 130 nm; (b) the liposome composition has a polydispersity index of less than 0.15, or the liposome composition has a polydispersity index of less than 0.10; (c) the drug / lipid ratio of AKG-28 to total phospholipid (PhL) in the composition is 99 to 530 g / mol PhL, or 85 to 456 g / mol as AKG-28 free base (FB); or AKG-28 free base (FB) of 85 to 400 g / mol; or 230 to 280 g / mol, or 190 to 240 g / mol as AKG-28 free base (FB); or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 290 to 360 g / mol or 245 to 305 g / mol as FB; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 300 to 340 g / mol or 256 to 290 g / mol as FB; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 250 g / mol or 215 g / mol as FB; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 250 g / mol or 215 g / mol as FB; or (d) the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 330 g / mol or 280 g / mol as FB; (e) the total concentration of AKG-28 in the composition is 8 to 15 mg / mL or 6.8 to 12.8 mg / mL as FB; or the total concentration of AKG-28 in the composition is 9 to 11 mg / mL or 7.6 to 9.4 mg / mL as FB; or the total concentration of AKG-28 in the composition is 10 mg / mL or 8.5 mg / mL as FB; or the total concentration of AKG-28 in the composition is 10 mg / mL (100 mg in a 10 ml vial) or 8 mg / mL as FB.(e) the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 90%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 95%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 97%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 98%; (f) the liposome composition comprises an aqueous medium containing sodium chloride and optionally containing ammonium ions; (g) the aqueous medium has an osmotic pressure of 270 to 330 mOsmol / kg; or the aqueous medium has an osmotic pressure of 270 to 310 mOsmol / kg; or the aqueous medium is isotonic; (h) the aqueous medium contains ammonium ions at a concentration of 20 to 60 mM; or the aqueous medium contains ammonium ions at a concentration of 50 to 80 mM; or the aqueous medium is 0 or the aqueous medium contains ammonium ions at a concentration of less than 0.5 mM; or the aqueous medium contains ammonium ions at a concentration of less than 1 mM; or the aqueous medium contains ammonium ions at a concentration of 1 to 10 mM; or the aqueous medium contains ammonium ions at a concentration of less than 130 mM; (i) the aqueous medium further contains a buffer, and the buffer buffers the liposome composition at a pH of 7.3 to 7.7 or a pH of 7.5; (j) the aqueous medium further contains HEPES or a phosphate buffer; or the aqueous medium contains HEPES at a concentration of 5 to 50 mM. (k) the composition further comprises a chelating agent; or the composition further comprises a chelating agent at a concentration of 0.1 to 1 mM; or the composition further comprises a chelating agent at a concentration of 0.5 mM; or the composition further comprises deferoxamine (DFO) or EDTA; or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM; or the composition further comprises 0.(1) further comprising deferoxamine (DFO) or EDTA at a concentration of 5 mM; or (2) the AKG-28 is encapsulated in the liposome as the sulfate salt of AKG-28.

[0025] One aspect of the present disclosure relates to a method for making a liposome composition, the method comprising the steps of: (a) dissolving one or more phospholipids, cholesterol, and a PEG-lipid derivative in ethanol to obtain a lipid solution; (b) combining the lipid solution of step (a) with a sequestering agent solution to obtain a homogenous lipid suspension having a desired phospholipid concentration; (c) extruding the lipid suspension of step (b) through a membrane having a defined pore size, such as a polycarbonate track-etched (PCTE) membrane having a nominal pore size of 50-200 nm; and (d) separating the lipids in the extruded lipid suspension. (e) contacting the liposomes with a compound of formula (I) in an aqueous medium to effect encapsulation of the compound in the liposomes; (f) optionally removing any unencapsulated compound; and (g) providing the liposomes in a physiologically acceptable medium suitable for parenteral use, wherein the sequestering agent solution of step (b) comprises an aqueous ammonium sulfate solution at a concentration greater than 0.25 M, and the physiologically acceptable medium of step (g) comprises a chelating agent.

[0026] In some embodiments, the capture agent solution in step (b) comprises ammonium sulfate at a concentration of 0.5 M. In some embodiments, the chelating agent is deferoxamine. In some embodiments, the liposomes extruded in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm. In some embodiments, the chelating agent is deferoxamine and the liposomes extruded in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm.

[0027] One aspect of the present disclosure relates to a method of treating a mycobacterial infection, the method comprising administering to a subject in need thereof a therapeutically effective amount of a liposome composition. In some embodiments, the mycobacterial infection is an infection with Mycobacterium tuberculosis, or an infection with a multidrug-resistant (MDR) strain of Mycobacterium tuberculosis, or an infection with an extremely drug-resistant (XDR) strain of Mycobacterium tuberculosis.

[0028] Provided herein is the use of liposomal compositions for treating mycobacterial infections.

[0029] Provided herein is the use of a liposomal composition for treating a methicillin-resistant Staphylococcus aureus (MRSA) bacterial infection.

[0030] In some embodiments, liposome preparations of oxazolidinone compounds are provided that have improved storage stability. In some embodiments, oxazolidinone liposome compositions containing more than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipids in the liposome composition and having a pH of 7 or greater have surprisingly improved storage stability. In some embodiments, the addition of a chelating agent such as deferoxamine or EDTA reduced oxidative degradation of cholesterol during storage of the oxazolidinone liposome composition. In some embodiments, oxazolidinone liposome compositions containing ammonium sulfate sequestered from liposomes containing an ammonium sulfate sequestering agent during oxazolidinone drug loading (e.g., by omitting buffer exchange after loading) showed improved phosphatidylcholine storage stability.

[0031] In some embodiments, the oxazolidinone liposome compositions provided herein comprise lipids consisting of HSPC, cholesterol, and PEG-DSPE in a mass ratio of about 5:3:1. In some embodiments, the oxazolidinone liposome compositions provided herein comprise lipids consisting of HSPC, cholesterol, and PEG-DSPE in a molar ratio of 45:55:2.25. In some embodiments, the oxazolidinone liposome compositions comprise an oxazolidinone consisting of AKG-28 or a pharmaceutically acceptable salt thereof. In some embodiments, the oxazolidinone liposome compositions comprise an oxazolidinone consisting of AKG-38 or a pharmaceutically acceptable salt thereof.

[0032] In some embodiments, a liposome composition is provided comprising a liposome vesicle and an oxazolidinone. In some embodiments, an oxazolidinone liposome composition is provided comprising a liposome vesicle encapsulating an oxazolidinone sulfate. In some embodiments, the liposome vesicle is in an aqueous medium.

[0033] In some embodiments, oxazolidinone compounds and their salts can be used to prepare liposome compositions. In some embodiments, the oxazolidinone liposome composition can be obtained by a process comprising combining an oxazolidinone compound with a purified, extruded lipid suspension under conditions effective to form oxazolidinone liposomes. The purified, extruded lipid suspension can comprise a lipid component consisting of phospholipids, cholesterol, and optionally a PEG-lipid derivative and a sequestering agent such as ammonium sulfate (AS) combined in an aqueous medium at a desired concentration. In some embodiments, the lipid component of the extruded lipid suspension consists of HSPC, cholesterol, and PEG(2000)-DSPE. In some embodiments, the lipid component of the extruded lipid suspension comprises HSPC and cholesterol in a molar ratio of 45:55. In some embodiments, the lipid component of the extruded lipid suspension comprises HSPC and cholesterol in a weight ratio of 5:3. In some embodiments, the lipid component of the extruded lipid suspension consists of HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25. In some embodiments, the lipid component of the extruded lipid suspension consists of HSPC, cholesterol, and PEG(2000)-DSPE in a weight ratio of 5:3:1.

[0034] In some embodiments, the purified extruded lipid suspension is obtained by a process comprising: (a) dissolving one or more phospholipids, cholesterol, and a PEG-lipid derivative in ethanol; (b) combining the lipid solution from step (a) with a sequestering agent solution (e.g., 0.5 M ammonium sulfate) to obtain a homogeneous lipid suspension having a desired phospholipid concentration (e.g., 60 mM phospholipid); (c) extruding the lipid suspension from step (b) through a membrane with a defined pore size, such as a polycarbonate track-etched (PCTE) membrane with a nominal pore size of 50-200 nm; and (d) purifying the liposomes from the extraliposomal sequestering agent in the extruded lipid suspension (e.g., by tangential flow filtration in a hollow fiber cartridge) to obtain a purified extruded liposome preparation. In some embodiments, the liposomes are mono- or oligolamellar vesicles with a z-average diameter of 90-130 nm or 100-130 nm. In some embodiments, the liposomes are mono- or oligolamellar vesicles with a polydispersity index of less than 0.15 or less than 0.10.

[0035] The purified extruded liposomes can be loaded with an oxazolidinone drug in a subsequent drug loading step. A drug stock of the oxazolidinone drug compound or salt thereof, at the desired drug and phospholipid concentrations, can be combined with the purified, extruded lipid suspension from step (d) to form a drug-liposome mixture under conditions effective to load the drug into liposomes in the purified, extruded liposome preparation. In some embodiments, the drug loading step involves exchange of a sequestering agent ammonium cation with the oxazolidinone compound across the liposome bilayer membrane, resulting in the production of extraliposomal ammonium in the drug-liposome mixture, which is displaced from inside the liposomes during the drug loading process.

[0036] After the drug loading process, the unencapsulated drug compound can be purified from the drug-liposome mixture (e.g., by size exclusion chromatography, SEC, dialysis, or diafiltration such as tangential flow filtration), and the composition comprising the oxazolidinone drug liposomes can be isolated and stored.

[0037] In some embodiments, the compound is entrapped in the liposome vesicle with a sequestering agent, and the sequestering agent comprises a polyanion. In some embodiments, the sequestering agent is triethylammonium sucrose octasulfate or ammonium sulfate. In some embodiments, the sequestering agent is triethylammonium sucrose octasulfate. In some embodiments, the sequestering agent is ammonium sulfate.

[0038] In some embodiments, the liposome composition comprises a salt of the compound, the salt being a sulfate, citrate, sucrosofate, a salt with a phosphorylated or sulfated polyol, or a salt with a phosphorylated or sulfated polyanionic polymer. In some embodiments, the liposome composition comprises a sulfate salt of the compound. For example, in some embodiments, the liposome composition comprises a sulfate or hydrosulfate salt of an oxazolidinone compound of Formula (I). In some embodiments, the liposome composition comprises a sulfate or hydrosulfate salt of (AKG-28). In some embodiments, the liposome composition comprises a sulfate or hydrosulfate salt of (AKG-38).

[0039] In some embodiments, the compound in the liposome vesicle has an aqueous solubility of less than 1 mg / mL. In some embodiments, the compound in the liposome vesicle has an aqueous solubility of less than 0.1 mg / mL.

[0040] In some embodiments, the liposome vesicle comprises a membrane comprising phosphatidylcholine and cholesterol. In some embodiments, the liposome vesicle comprises a membrane comprising phosphatidylcholine and cholesterol, the membrane separating the interior of the liposome vesicle from the aqueous medium. In some embodiments, the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soybean phosphatidylcholine (HSPC). In some embodiments, the molar ratio of phosphatidylcholine to cholesterol is about 60:40 to about 35:65. In some embodiments, the molar ratio of phosphatidylcholine to cholesterol is about 55:45 to about 35:65. In some embodiments, the molar ratio of phosphatidylcholine to cholesterol is about 50:50 to about 40:60. In some embodiments, the molar ratio of phosphatidylcholine to cholesterol is about 50:50 to about 45:55. In some embodiments, the membrane further comprises a polymer-conjugated lipid. In some embodiments, the liposome vesicles comprise HSPC, cholesterol, and polymer-conjugated lipids in a molar ratio of about 45:55:2.75. In some embodiments, the liposome vesicles comprise HSPC, cholesterol, and polymer-conjugated lipids in a molar ratio of 45:55:2.25. In some embodiments, the polymer-conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE). In some embodiments, the liposomes in the liposome composition have a Z-average particle size of about 80 to about 130 nm.

[0041] In some embodiments, the drug liposomes are provided in an aqueous medium containing sodium chloride, optionally further containing ammonium displaced from the liposomes during the drug loading process. In some embodiments, the concentration of sodium chloride in the liposome composition is 50-80 mM. In some embodiments, the concentration of sodium chloride in the liposome aqueous composition is 130-150 mM. In some embodiments, the drug liposomes are provided in an aqueous medium containing 20-60 mM ammonium displaced from the liposomes during the drug loading process. In some embodiments, the concentration of ammonium in the liposome aqueous medium is less than 0.5 mM. In some embodiments, the osmolality of the aqueous medium of the liposome composition is 270-330 mOsmol / kg. In some embodiments, the osmolality of the aqueous medium of the liposome composition is 270-310 mOsmol / kg.

[0042] In some embodiments, the oxazolidinone liposome composition has a pH greater than about 6.7. In some embodiments, the oxazolidinone liposome composition has a pH of 7-8. In some embodiments, the oxazolidinone liposome composition further comprises a buffer for adjusting the pH of the liposome aqueous medium to about 7.3-7.7. In some embodiments, the oxazolidinone liposome composition further comprises a buffer for adjusting the pH of the liposome aqueous medium to about 7.5. In some embodiments, the oxazolidinone liposome composition comprises a buffer substance selected from the group consisting of HEPES and phosphate. In some embodiments, the oxazolidinone liposome composition comprises a HEPES buffer. In some embodiments, the oxazolidinone liposome composition comprises a phosphate buffer. In some embodiments, the oxazolidinone liposome composition comprises a buffer substance selected from the group consisting of HEPES and phosphate at a concentration of 5-50 mM. In some embodiments, the oxazolidinone liposome composition comprises a buffer substance selected from the group consisting of HEPES and phosphate at a concentration of 20 mM.

[0043] In some embodiments, the oxazolidinone liposome composition further comprises a chelating agent. In some embodiments, the oxazolidinone liposome composition further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA. In some embodiments, the oxazolidinone liposome composition further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA at a concentration of 0.1 to 1 mM. In some embodiments, the oxazolidinone liposome composition further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA at a concentration of 0.5 mM.

[0044] In some embodiments, oxazolidinone drug compounds were efficiently loaded (>95%) into extruded liposomes at increased drug-to-lipid ratios (Example 51) with blood PK properties similar to those of liposomes with lower drug-to-lipid ratios (Example 53).

[0045] In some embodiments, oxazolidinone liposome preparations can be stabilized by retaining ammonium within the liposome that is displaced from the scavenger during the drug loading process (e.g., by omitting a buffer exchange step).

[0046] In some embodiments, the oxazolidinone drug compound is AKG-28, and the liposome composition comprises a sulfate salt of AKG-28 formed within the liposomes during the drug loading process. In some embodiments, the AKG-28 liposomes are prepared using a drug stock solution obtained by dissolving the salt form of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one (AKG-28). In some embodiments, a salt of AKG-28 is provided, including the hydrochloride salt of AKG-28. The salt of AKG-28 is useful for preparing a drug stock solution for loading AKG-28 liposomes. In some embodiments, AKG-28 ion-exchanges with ammonium displaced from the ammonium sulfate scavenger within the liposomes, forming an AKG-28 salt within the AKG-28 liposomes. [Table 3]

[0047] In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of AKG-28 of 230 to 380 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of 230 to 290 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of 290 to 360 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of 300 to 340 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of about 250 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has a drug / lipid ratio of about 330 g / mol total phospholipid (PhL). In some embodiments, the total (or aggregate) concentration of AKG-28 in the liposome composition is 8-15 mg / ml. In some embodiments, the total concentration of AKG-28 in the liposome composition is 9-11 mg / ml. In some embodiments, the ratio of encapsulated AKG-28 to total AKG-28 in the AKG-28 liposome composition is at least 90%, at least 95%, at least 97%, or at least 98%.

[0048] In some embodiments, the oxazolidinone drug compound is AKG-38, and the liposome composition comprises a sulfate salt of AKG-38 formed within the liposomes during the drug loading process. In some embodiments, the AKG-38 liposomes are prepared using a drug stock solution obtained by dissolving the salt form of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one (AKG-38). In some embodiments, a salt of AKG-38 is provided, including the hydrochloride salt of AKG-38. AKG-38 salts are useful for preparing drug stock solutions for loading AKG-38 liposomes. In some embodiments, AKG-38 ion-exchanges with ammonium displaced from the ammonium sulfate scavenger within the liposomes to form an AKG-38 salt within the AKG-38 liposomes. [Table 4]

[0049] In some embodiments, the AKG-38 liposome composition has AKG-38 at a drug / lipid ratio of 430 to 680 g / mol total phospholipid (PhL). In some embodiments, the AKG-38 liposome composition has AKG-38 at a drug / lipid ratio of 500 to 650 g / mol total phospholipid (PhL). In some embodiments, the AKG-28 liposome composition has AKG-38 at a drug / lipid ratio of 550 to 650 g / mol total phospholipid (PhL). In some embodiments, the AKG-38 liposome composition has AKG-38 at a drug / lipid ratio of about 450 g / mol total phospholipid (PhL). In some embodiments, the AKG-38 liposome composition has AKG-38 at a drug / lipid ratio of about 600 g / mol total phospholipid (PhL). In some embodiments, the total concentration of AKG-38 in the liposome composition is 12 to 25 mg / ml. In some embodiments, the total concentration of AKG-38 in the liposome composition is 13.5-16.5 mg / ml. In some embodiments, the total concentration of AKG-38 in the liposome composition is about 15 mg / ml. In some embodiments, the total concentration of AKG-38 in the liposome composition is about 20 mg / ml. In some embodiments, the ratio of encapsulated AKG-38 to total AKG-38 in the AKG-38 liposome composition is at least 90%, at least 95%, at least 97%, or at least 98%.

[0050] During accelerated stability testing of oxazolidinone liposome preparations, cholesterol and HSPC degradation was observed in certain AKG-28 and AKG-38 liposome compositions (Examples 54 and 65). Figure 17 is a scheme showing the two major cholesterol oxidative degradation products, 7-hydroxycholesterol (alpha and beta isomers) and 7-ketocholesterol. Figure 18 is a scheme showing the degradation of distearoylphosphatidylcholine (DSPC) to lysophosphatidylcholine (lyso-PC) and stearic acid. Hydrogenated soybean phosphatidylcholine (HSPC) is a 1,2-diacyl-sn-glycerophosphocholine with saturated fatty acids C16-C22 at the 1st and 2nd acyl chain positions, primarily stearic acid (C18) and palmitic acid (C16). Distearoylphosphatidylcholine is the largest component of HSPC.

[0051] However, in accelerated stability studies, in certain crude buffered liposomal formulations of AKG-28 prepared without a buffer exchange step, the formation of the HSPC degradation products lyso-PC, stearic acid, and palmitic acid was undetectable at a time when the buffer-exchanged formulations already exhibited HSPC degradation (Example 54). The extraliposomal medium in these formulations had increased levels of ammonium (substituted ammonium) displaced from the ammonium sulfate scavenger contained within the liposomal interior prior to the drug loading step. In some embodiments, AKG-28 liposomes may contain substituted ammonium in an amount equal to or exceeding the molar equivalent of the AKG-28 drug loaded into the liposomes. Similarly, HSPC degradation was minimized during accelerated stability testing in AKG-38 liposomes without buffer exchange after drug loading (Example 55).

[0052] Addition of the deferoxamine chelator to the liposome excipient buffer of AKG-28 and AKG-38 liposome compositions containing greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid surprisingly prevented cholesterol degradation throughout at least three months of accelerated stability testing at 37°C (Example 56). However, addition of DFO, EDTA, and DTPA chelators to the drug stock prior to drug loading did not protect against lipid degradation in AKG-28 and AKG-38 liposomes containing greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid (Example 57). Without the chelator (DFO or EDTA), oxidative degradation of cholesterol was observed in AKG-28 and AKG-38 liposome formulations (Examples 58 and 67).

[0053] In some embodiments, the liposome composition is stable against degradation of the liposomal lipid components and has a pH > 7.0. The rate of lipid degradation, particularly cholesterol degradation, was found to be dependent on the pH of the liposome formulation, being slower at pHs above 7.0 (Example 65). In some embodiments, the liposome composition has a pH of at least 7.1, at least 7.2, or at least 7.3, and a pH of 8.0 or less, pH 7.7 or less, or pH 7.6 or less. In some embodiments, the degree of cholesterol degradation after 3 months at 37°C is less than 10%, less than 5%, or less than 1% of total cholesterol. In some embodiments, the degree of phospholipid degradation after 6 weeks at 37°C is less than 10%, less than 5%, or less than 1% of total phospholipid. In some embodiments, the phospholipid is phosphatidylcholine. In some embodiments, the phospholipid is HSPC, and the pH is between 7.3 and 7.6.

[0054] In some embodiments, the liposome composition contains cholesterol and is stable against cholesterol degradation, and the degree of cholesterol degradation after 3 months at 37°C is less than 10%, less than 5%, or less than 1% of total cholesterol. It is important to avoid cholesterol degradation because the products of cholesterol degradation are toxic and can cause vascular endothelial damage (Rong et al., Arteriosclerosis, Thrombosis, and Vascular Biology, 1998, vol. 18, pp. 1885-1894; Sevanian et al., J Lipid Res, 1995, vol. 36, pp. 1971-1986). In some embodiments, the liposome composition contains a chelating agent. In some embodiments, the chelating agent is a chelating agent known to be well tolerated in humans. In some embodiments, the chelating agent is deferoxamine (desferal, DFO), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), including pharmaceutically acceptable salts thereof. In some embodiments, the chelating agent is present in the composition at a concentration of at least 0.01 mM, at least 0.05 mM, at least 0.1 mM, at least 0.2 mM, or at least 0.5 mM, and not more than 1 mM, 2 mM, 5 mM, or 10 mM. In some embodiments, the chelating agent is deferoxamine or deferoxamine mesylate, and the chelating agent concentration is about 0.5 mM.

[0055] In some embodiments, the external medium of the liposome composition has less than 0.5 mEq / L (milligram equivalents per liter) of ammonium or substituted ammonium. In some embodiments, the liposome composition contains ammonium or substituted ammonium in the external medium at a concentration of at least 10 mEq / L, at least 15 mEq / L, or at least 20 mEq / L, but not more than 200 mEq / L, not more than 150 mEq / L, not more than 100 mEq / L, not more than 80 mEq / L, or not more than 60 mEq / L. In some embodiments, the liposome composition contains ammonium or substituted ammonium in the extraliposomal medium at a concentration of at least 10 mEq / L, at least 15 mEq / L, or at least 20 mEq / L, but not more than 200 mEq / L, not more than 150 mEq / L, not more than 100 mEq / L, not more than 80 mEq / L, or not more than 60 mEq / L, and is stable against phospholipid degradation, with the extent of phospholipid degradation being less than 10%, less than 5%, or less than 1% of total phospholipid after 6 weeks at 37°C. In some embodiments, the phospholipid is phosphatidylcholine. In some embodiments, the phospholipid is HSPC, and the ammonium salt is ammonium chloride, ammonium sulfate, or a combination thereof, at an ammonium concentration of 10-80 mM or 15-60 mM. In some embodiments, the normality of ammonium in the external medium of the liposome composition is within 90-110% of the normality of the encapsulated drug in the drug loading step, where normality is the concentration expressed in gram equivalents / L (eq / L).

[0056] In some embodiments, the liposome composition comprises an encapsulated compound of Formula 1b at a drug / lipid (DL) ratio of 300-350 g / mol PhL. In some embodiments, the liposome composition comprises an encapsulated compound of Formula 1b at a DL ratio of 300-350 g / mol PhL and is characterized by an in vivo drug release half-life in the blood of CD-1 mice of greater than 80 hours, greater than 200 hours, or greater than 300 hours.

[0057] In some embodiments, the liposome composition comprises an encapsulated compound of Formula 1c at a DL ratio of 500-650 g / mol PhL.

[0058] In some embodiments, the liposome composition comprises liposomes in an aqueous medium, the liposomes being composed of HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1, the liposomes being mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm or 100-130 nm and a polydispersity index of less than 0.15 or less than 0.10, and the liposomes being 230-280 g / mol phospholipid (PhL), 290-360 g / mol PhL, 300-340 g / mol PhL, about 250 g / mol PhL, or about 330 g / mol PhL. The composition contains the encapsulated compound AKG-28 at a drug / lipid (DL) ratio of PhL, wherein the total concentration of AKG-28 in the composition is 8-15 mg / ml or 9-11 mg / ml, and the ratio of the encapsulated AKG-28 to the total AKG-28 in the composition is at least 90%, at least 95%, at least 97%, or at least 98%. In some embodiments, the aqueous medium contains sodium chloride and optionally ammonium ions. In some embodiments, the osmolality of the aqueous medium is 270-330 mOsmol / kg or 270-310 mOsmol / kg. In some embodiments, the ammonium concentration in the aqueous medium is 20-60 mM, and the sodium chloride concentration is 50-80 mM. In some embodiments, the ammonium concentration in the aqueous medium is less than 0.5 mM, and the sodium chloride concentration is 130-150 mM. In some embodiments, the composition also contains a buffering agent to adjust the pH of the aqueous medium to about 7.3-7.7 or about 7.5. In some embodiments, the buffering agent is HEPES or phosphate at a concentration of 5-50 mM or about 20 mM. The composition can also contain a chelating agent, which is deferoxamine (DFO) or EDTA at a concentration of 0.1-1 mM or about 0.5 mM. In some embodiments, the liposome composition is storage stable.

[0059] In some embodiments, the liposome composition comprises liposomes in an aqueous medium, the liposomes being composed of HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1, the liposomes being mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm or 100-130 nm and a polydispersity index of less than 0.15 or less than 0.10, the liposomes being 430-480 g / mol phospholipid (Phl), 500-650 g / mol Phl, 550-650 g / mol Phl, about 450 g / mol Phl, or about 600 g / mol Phl. The composition contains the encapsulated compound AKG-38 at a drug / lipid ratio of PhL, wherein the total concentration of AKG-38 in the composition is 12-25 mg / ml, 13.5-16.5 mg / ml, about 15 mg / ml, or about 20 mg / ml, and the ratio of encapsulated AKG-38 to total AKG-38 in the composition is at least 90%, at least 95%, at least 97%, or at least 98%. In some embodiments, the aqueous medium contains sodium chloride and optionally ammonium ions. In some embodiments, the osmolality of the aqueous medium is 270-330 mOsmol / kg or 270-310 mOsmol / kg. In some embodiments, the ammonium concentration in the aqueous medium is 20-60 mM, and the sodium chloride concentration is 50-80 mM. In some embodiments, the ammonium concentration in the aqueous medium is less than 0.5 mM, and the sodium chloride concentration is 130-150 mM. In some embodiments, the composition also contains a buffering agent to adjust the pH of the aqueous medium to about 7.3-7.7 or about 7.5. In some embodiments, the buffering agent is HEPES or phosphate at a concentration of 5-50 mM or about 20 mM. The composition can also contain a chelating agent, which is deferoxamine (DFO) or EDTA at a concentration of 0.1-1 mM or about 0.5 mM. In some embodiments, the liposome composition is storage stable.

[0060] In some embodiments, the liposome composition is stable to degradation of the encapsulated compound during storage. In some embodiments, the degradation of the encapsulated compound during storage under accelerated degradation conditions (37°C) is less than 5%, less than 4%, less than 3%, less than 2%, or less than about 1% after 3 months of storage, as measured by a decrease in compound purity expressed in percentage points. In some embodiments, the degradation of the encapsulated compound during storage under accelerated degradation conditions (37°C) is less than 20%, less than 10%, or less than 5% after 3 months of storage, as measured by a decrease in the total concentration of intact compound in the liposome composition. In some embodiments, the encapsulated compound is AKG-28 or AKG-38. Thus, liposomal compositions of AKG-38 stored at 37°C for 3 months showed a significantly lower decrease in AKG-38 purity from 98.99% to 98.07% (0.92 percentage points) and a lower overall decrease in intact AKG-38 concentration from 19.9 mg / ml to 19.06 mg / ml (4.2% drop) (Example 68). [Brief explanation of the drawings]

[0061] [Figure 1] 1 is a graph showing the effect of pH on liposome loading of compounds AKG-3, AKG-5, and AKG-16. [Figure 2] Figure 2A shows the effect of the added drug-to-lipid (DL) ratio on the liposome payload, expressed as the post-loading drug-to-lipid ratio (DL), in grams of drug per mole of liposomal phospholipid (PhL). Figure 2B shows the effect of the DL ratio (drug-to-lipid input ratio) on the liposome loading efficiency, calculated as the percentage of the post-loading DL relative to DL. [Figure 3A]1 is a graph showing the encapsulation of compounds AKG-5 and AKG-16 into liposomes using 0.5 M ammonium sulfate as a trapping agent at different DL ratios. The effect of DL ratio on liposome payload for AKG-5 and AKG-16 is shown. [Figure 3B] 1 is a graph showing the encapsulation of compounds AKG-5 and AKG-16 into liposomes using 0.5 M ammonium sulfate as a trapping agent at different DL ratios. The effect of DL ratio on liposome loading efficiency for AKG-5 and AKG-16 is shown. [Figure 3C] 1 is a graph showing the encapsulation of compound AKG-3 into liposomes using 0.5 M ammonium sulfate as a trapping agent at different DL ratios. FIG. 2 shows the effect of DL ratio on liposome payload for AKG-3. [Figure 3D] Figure 1 is a graph showing the encapsulation of compound AKG-3 into liposomes using 0.5 M ammonium sulfate as a trapping agent at different DL ratios. Figure 2 shows the effect of DL ratio on liposome loading efficiency for AKG-3. [Figure 4] Figure 4 shows graphs illustrating the encapsulation of AKG-28 and AKG-38 using TEA-SOS and ammonium sulfate as trapping agents at different DL ratios. Figure 4A shows the effect of DL ratio on liposome payload. Figure 4B shows the effect of DL ratio on loading efficiency. [Figure 5A] 1 is a graph showing the dependence of rapid drug leakage from liposomes encapsulating the compound AKG-28 upon in vitro contact with mouse (labeled "mouse") or human (labeled "human") plasma, as described in Example 19. Liposomes contained 5 mol% PEG(2000)-DSPE (labeled "DSPE") or PEG-DSG (labeled "DSG"). Scavenger: 0.5 M ammonium sulfate (AS). [Figure 5B]1 is a graph showing the dependence of rapid drug leakage from liposomes encapsulating the compound AKG-38 upon in vitro contact with mouse (labeled "mouse") or human (labeled "human") plasma, as described in Example 19. Liposomes contained 5 mol% PEG(2000)-DSPE (labeled "DSPE") or PEG-DSG (labeled "DSG"). Scavenger: 0.5 M ammonium sulfate (AS). [Figure 5C] 1 is a graph showing the dependence of rapid drug leakage from liposomes encapsulating the compound AKG-28 upon in vitro contact with mouse (labeled "mouse") or human (labeled "human") plasma, as described in Example 19. Liposomes contained 5 mol% PEG(2000)-DSPE (labeled "DSPE") or PEG-DSG (labeled "DSG"). Entrapment agent: 1N triethylammonium sucrose octasulfate (TEA-SOS). [Figure 5D] 1 is a graph showing the dependence of rapid drug leakage from liposomes encapsulating the compound AKG-38 upon in vitro contact with mouse (labeled "mouse") or human (labeled "human") plasma, as described in Example 19. Liposomes contained 5 mol% PEG(2000)-DSPE (labeled "DSPE") or PEG-DSG (labeled "DSG"). Entrapment agent: 1N triethylammonium sucrose octasulfate (TEA-SOS). [Figure 6] FIG. 1 depicts the numbered ring structures of compounds of formula (I). [Figure 7] Figure 1 shows the plasma concentration versus time profiles of total drug in Sprague-Dawley rats after administration of a single intravenous dose (IVx1) of Ls-AKG28 at 10 mg / kg (diamonds), 20 mg / kg (squares), and 40 mg / kg (circles). The plasma concentration versus time profiles of linezolid at 50 mg / kg (single oral dose, POx1) in 5% methylcellulose (pH 3-4) were also included for comparison. Mean and SD concentrations are presented at each time point. [Figure 8]Figure 1 shows the plasma concentration versus time profiles of total drug in Sprague-Dawley rats after administration of single intravenous doses (IVx1) of Ls-AKG38 at 20 mg / kg (diamonds), 40 mg / kg (squares), and 80 mg / kg (diamonds). The plasma concentration versus time profiles of linezolid at 50 mg / kg (single oral dose, POx1) in 5% methylcellulose (pH 3-4) were also included for comparison. Mean and SD concentrations are presented at each time point. [Figure 9A] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 10 mg / kg Ls-AKG28, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 9B] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 20 mg / kg Ls-AKG28, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 9C] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 40 mg / kg Ls-AKG28, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 10A] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 20 mg / kg Ls-AKG38, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 10B] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 40 mg / kg Ls-AKG38, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 10C] 1 is a graph showing plasma concentration versus time profiles of total drug in Sprague-Dawley rats following administration of 80 mg / kg Ls-AKG38, IVx1 on days 1 (circles), 15 (squares), 29 (diamonds), and 43 (triangles). Mean and SD concentrations are presented at each time point. [Figure 11A] 1 shows the plasma concentration versus time profiles of both lipid (non-exchangeable DiIC18(3)-DS labeled) and drug for liposomal AKG-28 in CD-1 mice after a single intravenous injection. Mean and SD concentrations are presented at each time point. [Figure 11B] 1 shows the plasma concentration versus time profiles of both lipid (non-exchangeable DiIC18(3)-DS labeled) and drug for liposomal AKG-38 in CD-1 mice after a single intravenous injection. Mean and SD concentrations are presented at each time point. [Figure 11C] 1 shows the change in plasma drug-to-lipid ratio, a measure of the drug release rate from liposomes, for both Ls-AKG28 and Ls-AKG38 in CD-1 mice after a single intravenous injection. Mean and SD concentrations are presented at each time point. [Figure 12] 1 is a graph showing plasma drug concentrations, presented as % injected dose, of compared multiple formulations of liposomal AKG-28 and liposomal AKG-38, Ls-AKG28 and Ls-AKG38, after the first and fourth weekly doses. Mice were injected with the indicated doses and formulations once a week for a total of four injections. [Figure 13A] 1 is a graph showing the effect of increasing doses of Ls-AKG28 on body weight in female CD-1 mice over time. [Figure 13B] 1 is a graph showing the effect of increasing doses of Ls-AKG38 on body weight in female CD-1 mice over time. [Figure 13C-1]1 is a graph showing the effect of Ls-AKG28 and Ls-AKG38 in combination with BP or BPM on hematological (RBC, HTC, PLT, WBC) and blood biochemistry (ALT, AST) parameters in female CD-1 mice. [Figure 13C-2] 1 is a graph showing the effect of Ls-AKG28 and Ls-AKG38 in combination with BP or BPM on hematological (RBC, HTC, PLT, WBC) and blood biochemistry (ALT, AST) parameters in female CD-1 mice. [Figure 13D] 1 is a heat map showing the effect of Ls-AKG28 or Ls-AKG38 monotherapy on histopathological findings in female CD-1 mice. [Figure 14A] 1 is a graph showing the effect of Ls-AKG28 in combination with bedaquiline and pretomanid (BP) or bedaquiline, pretomanid, and moxifloxacin (BPM) on body weight in female CD-1 mice over time. [Figure 14B] 1 is a graph showing the effect of Ls-AKG38 in combination with BP or BPM on body weight in female CD-1 mice over time. [Figure 14C-1] 1 is a graph showing the effect of Ls-AKG28 and Ls-AKG38 in combination with BP or BPM on hematological (RBC, HTC, PLT, WBC) and blood biochemistry (ALT, AST) parameters in female CD-1 mice. [Figure 14C-2] 1 is a graph showing the effect of Ls-AKG28 and Ls-AKG38 in combination with BP or BPM on hematological (RBC, HTC, PLT, WBC) and blood biochemistry (ALT, AST) parameters in female CD-1 mice. [Figure 14D] 1 is a heat map showing the effect of Ls-AKG28 and Ls-AKG38 in combination with BP or BPM on histopathological findings in female CD-1 mice. [Figure 15A]Graph showing weight change over time in female CD-1 mice treated with Ls-AKG28 injected at 50 mg / kg twice a week (2qw) or 100 mg / kg once a week (1qw), alone or in combination with BP. [Figure 15B] 1 is a graph showing body weight changes in female CD-1 mice treated with Ls-AKG38 injected 2 qw at 100 mg / kg or 1 qw at 200 mg / kg alone or in combination with BP. [Figure 15C-1] Graphs showing hematological and blood biochemistry parameters in female CD-1 mice treated with Ls-AKG28 (50 mg / kg 2 qw or 100 mg / kg 1 qw) or Ls-AKG38 (100 mg / kg 2 qw or 200 mg / kg 1 qw) alone or in combination with BP. [Figure 15C-2] Graphs showing hematological and blood biochemistry parameters in female CD-1 mice treated with Ls-AKG28 (50 mg / kg 2 qw or 100 mg / kg 1 qw) or Ls-AKG38 (100 mg / kg 2 qw or 200 mg / kg 1 qw) alone or in combination with BP. [Figure 15D] 1 is a heat map showing histopathological results in female CD-1 mice treated with Ls-AKG28 (50 mg / kg 2 qw or 100 mg / kg 1 qw) or Ls-AKG38 (100 mg / kg 2 qw or 200 mg / kg 1 qw) alone or in combination with BP. [Figure 16A] 1 is a graph showing the effect of Ls-AKG28 on body weight in male Sprague-Dawley rats chronically treated over time for a total of 8 weeks. [Figure 16B] 1 is a graph showing the effect of Ls-AKG38 on body weight in male Sprague-Dawley rats chronically treated for a total of 8 weeks over time. [Figure 17]1 is a scheme showing the two major cholesterol oxidative breakdown products, 7-hydroxy-cholesterol (alpha and beta isomers), and 7-ketocholesterol. [Figure 18] This is a scheme showing the decomposition of distearoylphosphatidylcholine (DSPC) into lysophosphatidylcholine and stearic acid. Hydrogenated soy phosphatidylcholine (HSPC) is a 1,2-diacyl-sn-glycerophosphocholine with saturated fatty acids C16-C22 in the 1st and 2nd acyl chain positions, primarily stearic acid (C18) and palmitic acid (C16). Distearoylphosphatidylcholine is the largest component of HSPC. [Figure 19A] 1 is a graph showing data on cholesterol degradation for AKG-38 liposomal compositions over a 12 week period at room temperature. [Figure 19B] 1 is a graph showing data on cholesterol degradation for AKG-28 liposomal compositions over a 12 week period at room temperature. [Figure 20A] 1 shows plasma concentration versus time profiles of AKG-28 drug, liposomal lipid (using a non-exchangeable DiIC18(3)-DS label), and plasma drug-to-lipid ratio for liposomal AKG-28 lots Ls-338 (Sample 71), Ls-339 (Sample 74), and Ls-340S (Sample 76) after a single intravenous injection in CD-1 mice. Liposome characteristics are shown in Example 59. Data points are the average of three animals. [Figure 20B] 1 shows plasma concentration versus time profiles of AKG-28 drug, liposomal lipid (using a non-exchangeable DiIC18(3)-DS label), and plasma drug-to-lipid ratio for liposomal AKG-28 lots Ls-338 (Sample 71), Ls-339 (Sample 74), and Ls-340S (Sample 76) after a single intravenous injection in CD-1 mice. Liposome characteristics are shown in Example 59. Data points are the average of three animals. [Figure 20C]1 shows plasma concentration versus time profiles of AKG-28 drug, liposomal lipid (using a non-exchangeable DiIC18(3)-DS label), and plasma drug-to-lipid ratio for liposomal AKG-28 lots Ls-338 (Sample 71), Ls-339 (Sample 74), and Ls-340S (Sample 76) after a single intravenous injection in CD-1 mice. Liposome characteristics are shown in Example 59. Data points are the average of three animals. [Figure 21] 1 is a graph showing data on cholesterol degradation for AKG-38 liposome composition lot Ls-371 (Example 67) when stored at 37° C. in the presence of various concentrations of deferoxamine. [Figure 22] 10 is a graph showing data on HSPC degradation of AKG-38 liposome composition lot Ls-371 (Example 67) when stored at 37° C. in the presence of various concentrations of deferoxamine. [Figure 23] 1 is a graph showing the pH change of AKG-38 liposome composition lot Ls-371 (Example 67) upon storage at 37° C. in the presence of various concentrations of deferoxamine. [Figure 24] 1 shows a synthetic scheme 1 according to one embodiment of the present disclosure. [Figure 25] 1 shows synthetic scheme 2 according to one embodiment of the present disclosure. [Figure 26] 1 shows synthetic scheme 3 according to one embodiment of the present disclosure. [Figure 27] 1 shows synthetic scheme 4 according to one embodiment of the present disclosure. [Figure 28] 1 shows synthetic scheme 5 according to one embodiment of the present disclosure. DETAILED DESCRIPTION OF THE INVENTION

[0062] The present disclosure describes oxazolidinone liposome compositions. In some embodiments, the liposome composition comprises a compound of Formula (I) encapsulated in lipid vesicles. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and cholesterol. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and more than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and more than about 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and 50-65 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and 50-60 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and 50-55 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and about 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition.In some embodiments, the liposome composition comprises a lipid vesicle comprising an oxazolidinone compound as a pharmaceutically acceptable salt thereof, a phospholipid, and about 55 mol % cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition.

[0063] In some embodiments, oxazolidinone liposome compositions are provided that are characterized by reduced phospholipid or cholesterol degradation during storage. In some embodiments, the oxazolidinone liposome compositions have a pH of about 7 or higher (e.g., 7-8) and contain phospholipids and greater than 50 mol% cholesterol (e.g., 50-65 mol%, 50-60 mol%, 50-55 mol%, about 50 mol%, or about 55 mol%) relative to the total cholesterol and non-PEGylated phospholipids in the liposome composition. In some embodiments, the oxazolidinone liposome compositions further contain a chelating agent, such as DFO or EDTA, in combination with the phospholipids and greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipids in the liposome composition. In some embodiments, the oxazolidinone liposome composition, having a pH of 7-8 (including 7-7.7, 7.1-7.7, 7.3-7.7, and about 7.5), further comprises a chelating agent, such as DFO or EDTA, in combination with a phospholipid and greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the oxazolidinone liposome composition further comprises extraliposomal ammonium in combination with a vesicle comprising a phospholipid and greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition. In some embodiments, the oxazolidinone liposome composition further comprises extraliposomal ammonium generated during drug loading of the oxazolidinone into liposome vesicles comprising a phospholipid and greater than 50 mol% cholesterol relative to the total cholesterol and non-PEGylated phospholipid in the liposome composition.

[0064] It is to be understood that the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the compositions and methods of the present disclosure.

[0065] [Definition] For convenience, certain terms employed in the specification, examples, and appended claims are collected here. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

[0066] As used herein, the following terms and phrases are intended to have the following meanings:

[0067] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0068] "Comprising" or "comprises" is used herein to refer to compositions, methods, and their respective component(s) that are present in a given embodiment but may include elements not specified.

[0069] "Consisting essentially of" is used herein to refer to elements required for a given embodiment. The term allows for the presence of additional elements that do not materially affect the basic and novel or functional characteristic(s) of that embodiment of the present disclosure.

[0070] "Consisting of" refers to compositions, methods, and each component thereof described herein, which excludes any element not recited in that description of an embodiment.

[0071] As used herein, "comprising" includes "consisting of" and "consisting essentially of."

[0072] When references are made herein to "as mentioned above" or "mentioned above" or "the foregoing", reference is made to any of the disclosures made herein in any of the preceding pages.

[0073] When references are made herein to "as referred to herein," "described herein," "provided herein," or "as referred to in this text," or "described herein," the specification refers to any of the disclosures made in the specification, either on the preceding or following pages.

[0074] "About" as used herein refers to an acceptable variation of within 20%, within 10%, and within 5% of the stated value. In certain embodiments, "about" can refer to a variation of ±1%, 2%, 3%, 4%, 5%, 10%, or 20%.

[0075] "Effective amount," as used herein with respect to a compound or composition, means an amount of an active compound (also referred to herein as an active agent or drug) sufficient to produce a bactericidal or bacteriostatic effect. In one embodiment, the effective amount is a "therapeutically effective amount," meaning the amount of active compound sufficient to alleviate the symptoms of the bacterial infection being treated.

[0076] A "subject" (or alternatively, a "patient"), as used herein, refers to an animal, preferably a mammal, most preferably a human, who receives prophylactic or therapeutic treatment.

[0077] "Administration" or "administering," as used herein, includes all means of introducing a compound or pharmaceutical composition into a subject in need thereof, including, but not limited to, oral, intravenous, intramuscular, intraperitoneal, subcutaneous, transdermal, inhalation, buccal, ocular, sublingual, vaginal, rectal, etc. Administration of the compound or composition is suitably parenteral. For example, the compound or composition may be administered preferentially intravenously, but may also be administered intraperitoneally or by inhalation, etc., currently in clinical use for liposomal amikacin in the treatment of Mycobacterium avium Complex Lung Disease. Drugs. 2019 Apr; 79(5):555-562.

[0078] "Treat", "treating", and "treatment" are used herein to refer to therapeutic or prophylactic measures such as those described herein.

[0079] "Synergistic" and "synergistic," as used herein, mean that the effect achieved with compounds used together is greater than the sum of the effects resulting from using the compounds separately, i.e., greater than the effect that would be predicted based on the two active ingredients administered separately.

[0080] "Pharmaceutically acceptable salt" refers to a relatively non-toxic, inorganic or organic acid addition salt of a compound according to the present disclosure, which possesses the desired pharmacological activity.

[0081] "Alkyl" means saturated carbon chains which may be linear or branched or combinations thereof, unless the carbon chain is defined otherwise. Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, sec- and tert-butyl, pentyl, hexyl, heptyl, octyl, and the like.

[0082] "Aminoalkyl" means an alkyl in which at least one carbon of the alkyl carbon chain forms a bond with an amino group, which may be a primary amino group, a monoalkyl-substituted (secondary) amino group, a dialkyl-substituted (tertiary) amino group, or an alkyl-substituted amino group, and the amine nitrogen atom and the alkyl chain replacing the amine hydrogen atom form a heterocyclic ring.

[0083] "Liposome" refers to a vesicle composed of a bilayer (unilamellar) and / or multiple concentric bilayers (multilamellar) separated by aqueous compartments formed by amphiphilic molecules such as phospholipids that surround a central aqueous compartment. In liposomal drug products, the active pharmaceutical ingredient is generally contained in a liposome. Typically, the water-soluble drug is contained in the aqueous compartment(s), and the hydrophobic drug is contained in the lipid bilayer(s) of the liposome. Drug release from liposomal formulations, particularly liposome clearance and other characteristics such as circulation half-life, can be modified by the presence of polyethylene glycol and / or cholesterol or other potential additives in the liposome.

[0084] A "unilamellar liposome," also known as a "unilamellar vesicle," is a liposome containing a single lipid bilayer membrane that defines a single, closed aqueous compartment. The bilayer membrane contains two layers of lipids: an inner layer and an outer layer (leaflet). The lipid molecules in the outer layer are oriented with their hydrophilic ("head") portions facing toward the outer aqueous environment and their hydrophobic ("tail") portions pointing downward toward the interior of the liposome. The inner lipid layer is located directly beneath the outer lipid layer, with the lipid heads facing the aqueous interior of the liposome and the lipid tails pointing toward the tails of the outer lipid layer.

[0085] "Multilamellar liposomes," also known as "multilamellar vesicles" or "multilamellar vesicles," contain more than one lipid bilayer membrane, which defines multiple closed aqueous compartments. The membranes are arranged concentrically, with different membranes separated by aqueous compartments.

[0086] "Encapsulated" and "entrapped," as used herein, refer to the incorporation or association of an oxazolidinone pharmaceutical agent in or with a liposome.

[0087] "DL," "DL ratio," "D / L," or "D / L ratio" are used interchangeably and refer to the ratio of drug to liposomal lipid. Unless otherwise specified, it is expressed as grams of drug per mole of liposomal phospholipid (PhL).

[0088] With respect to cholesterol, "mol%" refers to the molar amount of cholesterol relative to the sum of the molar amounts of cholesterol and non-PEGylated phospholipid, expressed in percentage points. For example, "55 mol% cholesterol" in a liposome containing cholesterol and HSPC refers to a composition of 55 mol parts cholesterol per 45 mol parts HSPC.

[0089] With respect to PEG-lipid, "mol%" refers to the ratio of the molar amounts of PEG-lipid to non-PEGylated phospholipid, expressed in percentage points. For example, "5 mol% PEG-DSPE" in a liposome containing HSPC and PEG-DSPE refers to a composition having 5 mol parts of PEG-DSPE per 100 mol parts of HSPC.

[0090] "Sucrose octasulfate," "sucrosofate," and "sucrooctasulfate" refer to the same compound, sucrose octasulfuric acid, or its anion, and are used interchangeably herein.

[0091] "Ac," "Me," and "Et" refer to the acetyl group (CH3CO), the methyl group (CH3), and the ethyl group (C2H5), respectively, in chemical formulas.

[0092] The term "free base concentration" or "FB concentration" is used to represent the mass concentration of a salt-forming compound in its free base form. By default, for compounds synthesized and isolated in salt form (e.g., the hydrochloride salt of a dihydrochloride salt), mass-based concentrations or ratios (e.g., mg / ml or g / mol phospholipid) are expressed as the concentration or ratio of this salt form. However, when written, the concentration of a compound isolated in salt form (e.g., AKG-28 dihydrochloride) is also expressed as the equivalent concentration of the compound as anhydrous free base (FB concentration). To obtain the FB concentration, the calculated molecular weight of the compound in free base form is divided by the calculated molecular weight of the salt form, and the concentration is multiplied by this factor. For example, the molecular weight of AKG-28 as a free base is 426.46, and the molecular weight of the dihydrochloride salt form (from which this compound is isolated) is 499.37. Therefore, the coefficient for calculating the concentration of AKG-28 on a FB basis is 426.46 / 499.37=0.854, so for example, 10 mg / ml of AKG-28 dihydrochloride has an FB concentration of 8.54 mg / ml. If the compound also contains a known amount of water (e.g., AKG-28 dihydrochloride monohydrate), a correction is also made for the known water content. The mass concentration of a compound isolated as the free base (e.g., AKG-38) is always expressed as the FB concentration.

[0093] Although the mass concentration and ratio disclosed herein depend on the salt form of compound, the concentration and ratio of compound can be expressed in molar units regardless of their salt form.For example, to convert the mass concentration or amount of AKG-28 into molar concentration as referred to herein based on its isolated synthetic product form of dihydrochloride, the mass concentration or amount is divided by the molecular weight of AKG-28 dihydrochloride, which is 499.4 g / mol.Therefore, the DL ratio of AKG-28 in liposome composition referred to herein as 330 g / mol PhL is expressed as 330 / 499.4=0.661 mol / mol PhL; the concentration of AKG-28 referred to as 10 mg / mL is expressed as 10 / 499.4=20.0 mM.When the concentration is referred to based on compound free base, the mass amount and concentration are divided by the molecular weight of compound free base. Thus, the DL ratio of AKG-38 in a liposome composition, quoted herein as 600 g / mol PhL, is expressed as 600 / 468.5 = 1.281 mol / mol PhL, and AKG-38 is isolated from synthesis in the form of the free base having a molecular weight of 468.5. Thus, the quoted 20 mg / mL concentration of AKG-38 is expressed in molar terms as 20 / 468.5 = 42.7 mM.

[0094] Various aspects and embodiments are described in detail in the subsections below.

[0095] [Compound] In some embodiments, a liposome composition comprising an oxazolidinone compound is provided.

[0096] Oxazolidinones are synthetic antibiotics that function by inhibiting protein synthesis. Linezolid (LZD) is an oxazolidinone compound that exhibits bacteriostatic activity against Mycobacterium tuberculosis. However, administration of LZD can cause serious side effects, such as anemia, thrombocytopenia, and peripheral neuropathy. Tedizolid is an oxazolidinone compound that has been shown to inhibit Gram-positive bacteria. Side effects of tedizolid phosphate are similar but generally milder than those of linezolid. However, compared with the experience of long-term administration of linezolid, tedizolid phosphate has limited experience with long-term administration, such as that required for the treatment of tuberculosis.

[0097] One aspect of the present disclosure relates to compounds that are aminoalkyl derivatives of oxazolidinone (see Figure 6). In some embodiments, compounds have the following formula (I): [ka] [In the formula, R2 is an amine (NH2) or acetamide (NHCOCH3), R1 is a tetrazole ring substituted at the 2' position with an aminoalkyl. and pharmaceutically acceptable salts thereof.

[0098] In some embodiments, the aminoalkyl is dimethylaminoalkyl. In some embodiments, the aminoalkyl derivative of the oxazolidinone compound contains either an amine group or an acetamide group at the R2 position of the oxazolidinone ring and a dimethylaminoethyl group on the tetrazole ring.

[0099] In another embodiment, the compound of formula (I): [ka] [In the formula, R2 is an amine (NH2) or acetamide (NHCOCH3), R1 is a tetrazole ring substituted at 1' with an aminoalkyl. and pharmaceutically acceptable salts thereof.

[0100] Aminoalkyl derivatives of oxazolidinone compounds having the chemical structures in Table 1 below were synthesized as described in Example 1.

[0101] The compounds of the present disclosure can exist in a free form, for example, as a free base, a free acid, or a zwitterion, or in the form of a salt. The salt can be any salt, organic or inorganic addition salt, or co-crystal commonly used in pharmacy, particularly any pharmaceutically acceptable organic or inorganic addition salt or co-crystal. It is understood that a chemical formula showing a particular salt or ionic form of a compound also discloses the undissociated free base (or free acid) form of the compound.

[0102] The present disclosure encompasses all stereoisomeric forms of the compounds. In some embodiments, the compounds in Table 1 below are substantially pure (i.e., at least 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, e.g., 100%).

[0103] [Table 5] JPEG2025501769000016.jpg244160JPEG2025501769000017.jpg240160JPEG2025501769000018.jpg158160

[0104] In some embodiments, the compound has the following chemical formula: [ka] It has.

[0105] In some embodiments, the compound has the following chemical formula: [ka] It has.

[0106] In some embodiments, the compound of Formula 1b is crystallized from aqueous ethanol. In some embodiments, the compound of Formula 1b is in the form of a dihydrochloride salt or a dihydrochloride monohydrate.

[0107] In some embodiments, the compound has the following chemical formula: [ka] It has.

[0108] In some embodiments, the compound has the following chemical formula: [ka] It has.

[0109] In some embodiments, the compound has the following chemical formula: [ka] It has.

[0110] Disclosed herein are compounds of formula (I) or pharmaceutically acceptable salts thereof that are useful for treating mycobacterial infections. In some embodiments, the compounds have formula 1a, 1b, 1c, 1d, or 1e. In some embodiments, the compounds have formula 1b: In some embodiments, the compound of formula (I) has a minimum inhibitory concentration (MIC), e.g., an MIC in the range of 0.1 μg / ml to 1 μg / ml, 0.25 μg / ml to 1 μg / ml, 0.5 μg / ml to 1 μg / ml, 0.1 μg / ml to 0.25 μg / ml, 0.1 μg / ml to 0.5 μg / ml, 0.25 μg / ml to 0.5 μg / ml, 0.01 μg / ml to 1 μg / ml, 0.01 μg / ml to 0.25 μg / ml, 0.01 μg / ml to 0.5 μg / ml, or 0.01 μg / ml to 0.1 μg / ml against Mycobacterium tuberculosis. In some embodiments, the compound of formula (I) has a minimum inhibitory concentration (MIC), e.g., an MIC of less than 1 μg / ml, less than 0.25 μg / ml, or less than 0.1 μg / ml against Mycobacterium tuberculosis. In some embodiments, the compounds of formula (I) have an MIC in the range of 0.01 μg / ml to 0.25 μg / ml. In some embodiments, the compounds of formula (I) have an MIC in the range of 0.01 μg / ml to 0.1 μg / ml. It should be understood that the MIC values ​​may be lower than the ranges provided herein, depending on the bacteria.

[0111] In some embodiments for treating mycobacteria, e.g., Mycobacterium tuberculosis, the compound (AKG-28 or AKG-38) has an MIC of less than 0.1 μg / mL. In some embodiments for treating mycobacteria, e.g., Mycobacterium tuberculosis, the compound has a selectivity index (SI) for killing Mycobacterium tuberculosis versus human kidney cells (VERO) of at least 1,000. In some embodiments for treating mycobacteria, e.g., Mycobacterium tuberculosis, the compound has an MIC of less than 0.1 μg / mL and a selectivity index (SI) for killing Mycobacterium tuberculosis versus human kidney cells (VERO) of at least 1,000. In some embodiments, the compound has the structure of AKG-28 (Formula 1b) or AKG-38 (Formula 1c). In some embodiments, the MIC is less than 0.05 μg / mL and the selectivity index for MIC compared to mitochondrial protein synthesis inhibition (SI-MPS) for Mycobacterium tuberculosis is greater than 20, such as AKG-28.

[0112] In some embodiments, the compounds described herein have a 2-20 fold increase (about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 20) in efficacy-adjusted dose compared to linezolid for Mycobacterium tuberculosis.

[0113] In some embodiments, for treating methicillin-resistant Staphylococcus aureus (MRSA), the compound has an MIC against MRSA strains of less than 2 μg / mL. In some embodiments for treating methicillin-resistant Staphylococcus aureus (MRSA), the compound has an IC50 against human VERO kidney cells of greater than 100 μg / mL. In some embodiments for treating methicillin-resistant Staphylococcus aureus (MRSA), the compound has an MIC against MRSA strains of less than 2 μg / mL and an IC50 against human VERO kidney cells of greater than 100 μg / mL. In some embodiments, the compound has the structure of AKG-38 (Formula 1c), AKG-39 (Formula 1e), and AKG-40 (Formula 1d).

[0114] (Water solubility) In some embodiments, the compound is in the form of a salt, such as a hydrochloride or mesylate, and is soluble in water at a concentration of more than 1 mg / ml, preferably more than 10 mg / ml (and up to 1 g / ml) before encapsulation in liposomes. Additional salts before encapsulation may include, but are not limited to, besylate, bitartrate, carbonate, citrate, esylate, gluconate, glutamate, glycolate, lactate, malate, maleate, mandelate, methylsulfate, napsylate, phosphate, propionate, salicylate, succinate, tartrate, and tosylate. In some embodiments, the compound is in the form of a hydrate, solvate, or co-crystal before encapsulation in liposomes.

[0115] In some embodiments, drugs are entrapped inside liposomes in different salt forms with reduced aqueous solubility, e.g., less than 1 mg / mL, preferably less than 0.1 mg / mL (0.1-0.001 mg / mL). Salts of compounds once entrapped in liposomes include, but are not limited to, sulfate, citrate, phosphate, sucrose, or various phosphorylated or sulfated polyols or polyanionic polymers. Exemplary polyols include, but are not limited to, sucrose, erythritol, mannitol, xylitol, sorbitol, inositol, and combinations thereof. Exemplary polyanionic polymers include, but are not limited to, polyvinyl sulfonate, polyvinyl sulfonate, polyphosphate, copolymers of acrylic acid and vinyl alcohol sulfate, and combinations thereof.

[0116] Working stocks of compounds were prepared as follows: To an aliquot of the compound (free base) in powder form, 1 to 1.5 equivalents of HCl in the form of a 1N aqueous solution was added, and the mixture was vortexed until homogeneous. To the resulting cake or syrup, water was added to a final concentration of typically 10 mg / ml, and complete dissolution was observed. In some cases, 0.95 equivalents of HCl were added to the free base form of the drug to prepare a 20 mg / ml stock solution.

[0117] The aqueous solubility of the compounds according to the present disclosure is illustrated by the following observations to obtain a visually clear solution.

[0118] [Table 6]

[0119] These results demonstrate that the compounds provided herein: -Linezolid (3mg / ml) (www.drugbank.ca / drugs / DB00601) [ka] -Stezolid (0.237mg / ml) (www.drugbank.ca / drugs / DB11905) [ka] and -Tedizolid (0.382 mg / mL) (www.drugbank.ca / drugs / DB14569) [ka] This indicates that the compound has a higher water solubility than the known water solubility of

[0120] In some embodiments, prior to encapsulation in liposomes, the aqueous solubility of the compounds described herein is at least 5 times, at least 10 times, at least 20 times, at least 30 times, or at least 40 times that of the oxazolidinone.

[0121] The excellent water solubility of the compounds described herein, along with their amphiphilic weak base properties, allows for the efficient use of transmembrane gradient-based and intraliposomal complexation (active loading) approaches to generate liposome-encapsulated forms of these compounds with high drug / carrier (drug / lipid) ratios and pharmacokinetic properties favorable for delivery of the encapsulated drug to infected tissues after systemic drug administration. As used herein, an amphiphilic weak base has a pKa of 7-12 and a logP of 1-6.

[0122] Liposome loading properties and antimycobacterial activity An important feature of the compounds described herein is their weakly amphipathic basic character, which facilitates transmembrane gradient-driven loading of these compounds into liposomes. In some embodiments, the weakly basic character of the compounds disclosed herein is characterized by an electrolytic dissociation constant in the pKa range of 7.0-12.0, 7.5-11.0, 7.8-10.5, or 8.0-10.0. In some embodiments, the amphipathic character of the compounds described herein is characterized by a logP parameter in the range of 0.5-5.0, 1.0-4.0, 1.0-3.5, or 1.0-3.0. It has been unexpectedly discovered that certain embodiments possessing these favorable properties for liposome loading also possess superior activity against mycobacteria, matching or exceeding the activity of similar compounds in the same class of drugs that have unfavorable properties for efficient and stable liposome encapsulation.

[0123] (Liposome composition) Compositions and uses of the compositions for treating tuberculosis and other mycobacterial and gram-positive bacterial infections are disclosed. The compositions provided herein contain highly potent, selective oxazolidinones that are encapsulated with high efficiency to maximize the drug's dosability and are stable in the presence of plasma. In some embodiments, the compositions are long-circulating and retain their encapsulated drug in the circulation after intravenous administration, allowing for efficient accumulation at the site of bacterial or mycobacterial infection. In some embodiments, the high doses that can be achieved, combined with the long-circulating properties and highly stable drug retention, allow for reduced dosing frequency compared to the once-daily or twice-daily administration of other drugs typically used to treat these infections.

[0124] Disclosed herein are pharmaceutical compositions for treating bacterial infections, particularly Mycobacterium tuberculosis infections. In some embodiments, the pharmaceutical composition is a liposome composition comprising a polyanion or a sulfate salt containing a polyanion and an aminoalkyloxazolidinone compound.

[0125] Another aspect of the present disclosure relates to a method of treating a bacterial infection, the method comprising administering to a subject in need thereof a therapeutically effective amount of a liposome composition provided herein.

[0126] In some embodiments, the bacterial infection is a Mycobacterium tuberculosis infection. In some embodiments, the compound in the liposome vesicle has a minimum inhibitory concentration (MIC) in the range of about 0.01 μg / ml to about 0.25 μg / ml. In some embodiments, the compound in the liposome vesicle has a minimum inhibitory concentration (MIC) in the range of about 0.01 μg / ml to about 0.1 μg / ml.

[0127] In some embodiments, the composition comprises liposomes in a medium, the intraliposomal space comprising an aqueous phase having a polyanion and a compound of Formula (I). In some embodiments, the composition comprises liposomes in a medium, the intraliposomal space comprising a polyanion or a sulfate salt containing a polyanion and the compound AKG-16, AKG-28, or AKG-38. In some embodiments, the medium is an aqueous medium, and the primary composition in the medium is a compound of Formula (I) and a corresponding scavenger.

[0128] The compounds of formula (I) can be entrapped in liposomes using a suitable polyanion, such as sucrose octasulfate (e.g., derived from a triethylammonium sucrose octasulfate (TEA-SOS) gradient) or sulfate (e.g., derived from an ammonium sulfate gradient). Additional polyanion entrapment agents include, but are not limited to, inositol hexaphosphate, inositol hexasulfate, polyvinylsulfonic acid, dextran sulfate, citrate, polyphosphate, and suramin.

[0129] The external aqueous medium typically contains an appropriate buffer and isotonicity agent. Suitable buffers include histidine, citrate, HEPES, MOPS, MES, TRIS, phosphate, glycine, imidazole, borate, carbonate, and succinate. Isotonicity agents include salts such as sodium chloride, potassium chloride, sucrose, glycerin, dextrose, or mannitol.

[0130] In some embodiments, the composition comprises a compound of Formula (I), or Formula 1a, 1b, 1c, or 1d, or a pharmaceutically acceptable salt thereof, encapsulated with a polyanion in a primarily unilamellar vesicle formed from one or more phospholipids, a sterol, and a lipid optionally conjugated to a hydrophilic polymer (polymer-conjugated lipid). In some embodiments, the composition can comprise a compound of Formula (I), or Formula 1a, 1b, 1c, or 1d, or a pharmaceutically acceptable salt thereof, encapsulated with a polyanion in unilamellar and multilamellar vesicles (e.g., having two or three layers). It should be understood that multilamellar vesicles may be cleared from circulation more rapidly than unilamellar vesicles. In some embodiments, the phospholipid is hydrogenated soybean phosphatidylcholine (HSPC), distearoylphosphatidylcholine (DSPC), or egg sphingomyelin (ESM). The term "phospholipid" as used herein refers to any one or combination of phospholipids that can form liposomes.Neutral phospholipids can include diacylphosphatidylcholine, dialkylphosphatidylcholine, sphingomyelin, and diacylphosphatidylethanolamine.Phosphatidylcholine (PC) includes those obtained from egg, soybean, or other plant sources, or those partially or completely synthetic, or those with variable lipid chain length and unsaturation, and are suitable for use in the present composition. Synthetic, semi-synthetic and natural product phosphatidylcholines, including but not limited to distearoylphosphatidylcholine (DSPC), hydrogenated soy phosphatidylcholine (HSPC), soy phosphatidylcholine (soy PC), egg phosphatidylcholine (egg PC), hydrogenated egg phosphatidylcholine (HEPC), dipalmitoylphosphatidylcholine (DPPC) and dimyristoylphosphatidylcholine (DMPC), are suitable phosphatidylcholines for use in the present disclosure.Charged phospholipids can include phosphatidylserine, phosphatidic acid, phosphatidylinositol, phosphatidylglycerol, cardiolipin, or head group-modified lipids such as N-succinyl-phosphatidylethanolamine, N-glutaryl-phosphatidylethanolamine, and PEG-derivatized phosphatidylethanolamine.

[0131] Polymer-conjugated lipids include poly(ethylene glycol)-conjugated (PEGylated) phospholipids (PEG-lipids), such as PEG(molecular weight 2,000) methoxy-poly(ethylene glycol)-1,2-distearoyl-sn-glycerol (PEG(2000)-distearoylglycerol, PEG-DSG), PEG(molecular weight 2,000) 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (PEG(molecular weight 2,000)-distearoylphosphatidylethanolamine, PEG-DSPE), or PEG(molecular weight 2,000) N-palmitoyl-sphingolipids. The molecular weight of the PEG moiety in the PEG-lipid component can also vary from 500 to 10,000 g / mol, 1,500 to 6,000 g / mol, but is preferably approximately 2,000 MW. Other polymers used for conjugation to lipid anchors include poly(2-methyl-2-oxazoline) (PMOZ), poly(2-ethyl-2-oxazoline) (PEOZ), poly-N-vinylpyrrolidone (PVP), polyglycerol, poly(hydroxyethyl L-asparagine) (PHEA), and poly(hydroxyethyl L-glutamine) (PHEG).

[0132] In some embodiments, the sterol is cholesterol. Other sterols include, but are not limited to, phytosterols, such as ergosterol and β-sitosterol, and hopanoids. In some embodiments, the ratio of phospholipid(s) to cholesterol is selected to provide a desired amount of liposome membrane rigidity while maintaining a sufficiently reduced amount of leakage of the compound of Formula (I) from the liposomes. In some embodiments, a polymer-conjugated lipid can be optionally added to reduce the tendency of the liposomes to aggregate. The type and amount of polymer-conjugated lipid can be selected to provide a desired level of protein binding, liposome stability, and circulation time in the bloodstream. For example, liposome vesicles contain phosphatidylcholine (e.g., DSPC or HSPC) and cholesterol in a molar ratio of about 45:55. The molar ratio of phosphatidylcholine to cholesterol can vary from about 60:40 to 35:65, from about 50:50 to 35:65, or from about 50:50 to about 45:55. In particular, liposomes can contain vesicles composed of HSPC, cholesterol, and polymer-conjugated lipids (PEG-DSG or PEG-DSPE) in a molar ratio of approximately 45:55:2.75, corresponding to a 5 mol% PEG-lipid concentration relative to the phospholipid concentration. The PEG-lipid concentration can vary from 0.5 to 10 mol% relative to the (non-PEGylated) phospholipid, with a preferred ratio being 3 to 10 mol%, and an even more preferred ratio being 4 to 8 mol%.

[0133] In some embodiments, the liposome composition provides desirable pharmacokinetic properties, such as an extended plasma half-life, measured as the percentage of the injected dose (ID) (or injected amount) remaining in the blood 6 or 24 hours after intravenous injection in immunocompetent mice, and allows stable encapsulation of the drug in the plasma over 24 hours, as determined by the change in the drug-to-lipid ratio (DL ratio) after iv administration in mice. In some embodiments, the percentage of drug remaining in the blood is greater than 20%, preferably greater than 30%, and most preferably greater than 40% of the injected dose at 6 hours. The percent retained in the blood after 24 hours is preferably greater than 10%, more preferably greater than 20% of the injected dose. The DL ratio is greater than 20% at 24 hours, preferably greater than 50%, and most preferably greater than 80% of the original injected liposomal drug. Desirable liposome compositions also exhibit stable encapsulation in the presence of human plasma in vitro using a burst release method, with the liposomes retaining greater than 50% of the drug over 20 minutes, and greater than 60%, greater than 70%, preferably greater than 80%, and most preferably greater than 90% of the encapsulated drug over 20 minutes.

[0134] The liposomes according to the present disclosure can be prepared by any method known in the art. For example, see G. Gregoriadis (editor), Liposome Technology, vol. st edition,1983; 2 nd edition,1993; 3 rdSee, e.g., "The Liposomes of the Present Disclosure," ed., 2006; CRC Press, Boca Raton, Fla. Examples of suitable methods for producing liposome compositions according to the present disclosure include membrane extrusion, reverse-phase evaporation, sonication, solvent (e.g., ethanol) injection (including microfluidics, Y-junction, and T-junction mixing), microfluidization, detergent dialysis, ether injection, and dehydration / rehydration. Liposome size can be controlled by controlling the pore size of the membrane used for extrusion, or the pressure and number of passes used in microfluidization or any other suitable method. In some embodiments, the desired lipids are first hydrated by thin-film hydration or ethanol injection, and then sized by extrusion through membranes of defined pore sizes, such as 50 nm, 80 nm, 100 nm, or 200 nm, or a combination thereof, to produce liposomes with an average size in the range of 70-150 nm, or 80-130 nm, and a polydispersity index of 0.1 or less. The drug compound to be encapsulated is added to liposomal lipid before liposome formation, and then dissolved in the aqueous medium that liposome is formed by the above-mentioned method, thereby isolating the drug in liposome.In some embodiments, drug compound is encapsulated in liposome by using a capture agent that is incorporated into the inner space of liposome (see Drummond, DC, et al. (2006) in: Liposome Technology, Third Edition (Ed. Gregoriadis, G.) Volume 2, p.149-168).

[0135] In some embodiments, a method for making a liposome composition according to the present disclosure includes the steps of: (i) preparing liposomes comprising phospholipids, cholesterol, and PEG-lipids, the liposomes having an interior space containing a sequestering agent in a medium substantially free of the sequestering agent; (ii) contacting the liposomes with a compound according to the present disclosure in an aqueous medium to encapsulate the compound in the liposomes; (iii) removing the unencapsulated compound; and (iv) providing liposomes in a physiologically acceptable medium suitable for parenteral use. In some embodiments, if the encapsulation efficiency of step (ii) is sufficiently high, typically >95%, >97%, or >99%, step (iii) of removing the unencapsulated compound is omitted.

[0136] In some embodiments, a method for forming liposomes with a compound therein includes (a) preparing liposomes containing a sequestering agent consisting of an ammonium or substituted ammonium salt of a polyanion; (b) subsequently removing the sequestering agent outside the liposomes to form an electrochemical gradient across the membrane; and (c) contacting the liposomes with the compound under conditions effective to allow the compound to enter the liposomes and a corresponding amount of ammonia or substituted ammonia to leave the liposomes (thereby depleting or reducing the resulting pH gradient across the liposomes). Liposome compositions containing a sequestering agent inside the liposomes can be produced by forming liposomes in a solution of the sequestering agent. A transmembrane concentration gradient of the sequestering agent can be formed across the liposomes by removing the sequestering agent outside the liposomes or by diluting the liposomes either after liposome formation or before drug loading (sequestering).

[0137] In some embodiments, the contacting step involves incubating the liposomes with the drug in an aqueous medium at a temperature above ambient temperature but below the boiling point of water, preferably between 30°C and 90°C, 40°C and 80°C, 50°C and 80°C, or 60°C and 75°C. In some embodiments, the incubation is performed at an ionic strength less than that corresponding to 50 mM NaCl, or more preferably less than that corresponding to 30 mM NaCl. After incubation, concentrated salt, e.g., NaCl solution, can be added to raise the ionic strength to above that of 50 mM NaCl, or about 100 mM NaCl. Increasing the ionic strength after the drug-loading incubation step helped reduce post-loading aggregation of the liposomes. Incubation times can range from several minutes to several hours. In some embodiments, the incubation time is 5 to 40 minutes, 10 to 30 minutes, or 15 to 25 minutes. After incubation, the liposomes are cooled and then allowed to reach ambient temperature. In some embodiments, liposomes are cooled to 2-15°C. In some embodiments, liposomes are cooled to 4-10°C. After the cooling step, a concentrated salt, e.g., NaCl, solution can be added to increase the ionic strength to greater than 50 mM NaCl, or about 100 mM NaCl. Increasing the ionic strength after the drug loading incubation step helped reduce aggregation after liposome loading.

[0138] In another embodiment, loading is performed at 20-350 mEq / L, 20-100 mEq / L, or 50-80 mEq / L in the presence of an ionic agent, such as NaCl, KCl, NH4Cl, Na2SO4, K2SO4, or (NH4)2SO4. Contrary to the art-wide practice of low ionic strength (low salt concentration), loading of compounds of the present disclosure, particularly AKG-28, into liposomes has been found to be more efficient in the presence of relatively high ionic strength agents, such as NaCl, particularly when loading is performed at higher drug concentrations. Thus, with 10-12 mg / ml of AKG-28 in the liposome loading mixture, loading at 0-20 mM NaCl resulted in encapsulation efficiencies of approximately 90-94%, while loading at approximately 80-360 mM NaCl resulted in loading efficiencies of >97% (Example 62). Thus, in some embodiments of the present disclosure, loading of the compounds described herein is performed at 20-350 mEq / L, 20-100 mEq / L, or 50-80 mEq / L of ionic strength agent. In some embodiments, the ionic strength agent is NaCl. In some embodiments, the concentration of the added ionic strength agent is selected so that the loaded liposomes are isotonic (having an osmolality of 280-310 mOsmol / L, or 270-310 mOsmol / kg). In some embodiments, the drug is AKG-28, the ionic strength agent is NaCl, and loading is performed at approximately 12-13 mg / ml of drug and 50-80 mM NaCl. Encapsulation efficiencies of 95% or greater, 97% or greater, or 98% or greater can be achieved.

[0139] In some embodiments, the contacting step also includes incubating the liposomes with the drug in an aqueous medium in the presence of an osmotic (tonicity) balancing agent. In some embodiments, the osmotic balancing agent (also referred to herein as an osmotic agent) is a non-ionic agent. Exemplary non-ionic osmotic agents include, but are not limited to, dextrose (glucose), sucrose, trehalose, lactose, mannitol, sorbitol, and polyvinylpyrrolidone. In some embodiments, the concentration of the osmotic agent has an osmolarity (expressed as osmolarity) equal to the osmolarity of the sequestering agent solution in the interior space of the liposome prior to drug loading. The osmolarity of the sequestering agent solution can be measured by methods known in the art before the solution is combined with lipids to form liposomes. In another embodiment, the concentration of the osmotic agent is lower than the osmotic concentration of the sequestering agent solution, providing an osmotic concentration that is less than about 90%, less than about 80%, less than about 70%, less than about 60%, less than about 50%, less than about 40%, less than about 30%, less than about 20%, or less than about 10% of the osmotic concentration of the sequestering agent solution. In yet another embodiment, the concentration of the osmotic agent during the drug loading process is in the range of 200-400 mmol / kg, preferably 250-350 mmol / kg. In yet another embodiment, the osmotic agent is dextrose at a concentration of 45 g / L. In yet another embodiment, no osmotic agent is used during the incubation of the liposomes with the drug. In yet another embodiment, the incubation is performed in the presence of an ionic strength adjuster. A non-limiting example of an ionic strength adjuster is sodium chloride, which is added to the liposome-drug solution at a concentration of, for example, 5-50 mM, 10-20 mM, or about 10 mM. Contrary to convention in the liposome field, compounds according to the present disclosure, such as AKG-28 and AKG-38, are loaded into liposomes according to the present disclosure in a stable and highly efficient manner, even when the amount of osmotic agent during the drug-liposome contacting step provides an osmotic concentration lower than that of the scavenger solution (osmotically unbalanced liposomes) until the complete absence of added osmotic agent.

[0140] For compounds AKG-28 and AKG-38, surprisingly, loading was found to be highly effective (>95% loading, >97% loading, and >98% loading) even at the upper limits of the achievable DL ratio (AKG-28, 300-350 g / mol PhL; AKG-38, 500-600 g / mol PhL) and at high drug concentrations in the liposome-drug loading mixture (>16 mg / ml for AKG-38 and >12 mg / ml for AKG-28). Typically, in some embodiments, liposome loading of AKG-28 is performed at 300-350 g / mol PhL with a drug concentration of 6 mg / ml, at least 10 mg / ml, or at least greater than 12 mg / ml, while liposome loading of AKG-38 is performed at 500-650 mg / ml, or 500-600 mg / ml of drug with a drug concentration of 8 mg / ml, at least 12 mg / ml, or at least greater than 16 mg / ml, achieving an efficiency of at least 95% loading, at least 97% loading, or at least 98% loading.

[0141] In some embodiments, compounds of the present disclosure are loaded into liposomes over a wide pH range, such as pH 4.5 to 7. For AKG-28, optimal loading efficiencies of 95% or greater, or even 97% or greater, were achieved in the pH range of 5.5 to 7.0 (Example 62). The loading pH is defined by the pH of the drug aqueous stock solution (40 mg / ml), selected from the pH range of 5.3 to 7.0. In some embodiments, the pH of the 40 mg / ml AKG-28 stock solution is in the pH range of 5.7 to 6.9, adjusted with NaOH.

[0142] Liposomes and other lipid nanoparticle compositions are susceptible to lipid component degradation during storage, thereby adversely affecting their pharmaceutical quality. Lipid degradation is generally assumed to follow the Arrhenius law, but can be studied in an accelerated stability study format, where liposome samples are stored at temperatures higher than the suggested storage temperature, so that degradation occurs more rapidly. For example, liposomes according to the present disclosure, containing compounds AKG-28 and AKG-38 in a lipid composition of PC and cholesterol, were found to accumulate both cholesterol oxidative degradation products (Figure 17) and degradation products from phosphatidylcholine hydrolysis (Figure 18). While oxidative degradation is typically observed in lipids with unsaturated hydrocarbon chains, oxidative degradation of cholesterol is an uncommon phenomenon. Stabilization of liposomes with the encapsulated compounds described herein can be achieved by the incorporation of chelating agents. Chelating agents are molecules that bind metal ions by forming one or more stable heterocyclic groups containing metal and coordinate bonds. Exemplary chelating agents are deferoxamine (desferrioxamine, desferal) (abbreviated herein as DFO), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). In some embodiments, the liposome composition comprises cholesterol and is stable against cholesterol degradation, with the degree of cholesterol degradation being less than 10%, less than 5%, or less than 1% of total cholesterol after 3 months at 37°C. In some embodiments, the liposome composition comprises a chelating agent.In some embodiments, the chelating agent is deferoxamine (desferal, DFO), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), including pharmaceutically acceptable salts thereof. The chelating agent can be present in the composition at a concentration of at least 0.01 mM, at least 0.05 mM, at least 0.1 mM, at least 0.2 mM, or at least 0.5 mM, and at most 1 mM, 2 mM, 5 mM, or 10 mM. In some embodiments, the chelating agent is deferoxamine or deferoxamine mesylate, and the chelating agent concentration is about 0.5 mM. Deferoxamine has been found to be particularly effective in preventing cholesterol degradation in liposomes according to the present disclosure. The stability of lipids with encapsulated compounds according to the present disclosure, particularly AKG-28 and AKG-38, was affected by the pH of the extraliposomal medium. While the general teaching in the field is that optimal lipid stability in liposomes is achieved at a pH of about 6.5, it has been discovered that optimal lipid stability for both cholesterol and PC components is achieved at a pH above 7.0 for liposomes according to the present disclosure. In some embodiments, the liposome composition has a pH of at least 7.1, at least 7.2, or at least 7.3, and is no greater than pH 8.0, no greater than pH 7.7, or no greater than pH 7.6. In some embodiments, the degree of cholesterol degradation after 3 months at 37°C is less than 10%, less than 5%, or less than 1% of total cholesterol. In some embodiments, the extent of phospholipid degradation after 6 weeks at 37°C is less than 10%, less than 5%, or less than 1% of the total phospholipid content. In some embodiments, the phospholipid is phosphatidylcholine. In some embodiments, the phospholipid is HSPC and the pH is 7.3-7.6.

[0143] Another factor that unexpectedly affected the stability of lipids in the liposomes of the present disclosure was the presence of ammonium or substituted ammonium in the external medium. In some embodiments, ammonium salts are used as sequestering agents to effect liposomal loading of compounds described herein, such as AKG-28 or AKG-38. Thus, for each molecule of drug that enters the liposome interior, one or two molecules of ammonia leave the liposome interior and accumulate in the extraliposomal medium. Subsequently, the accumulated ammonium is purged during a post-loading buffer exchange / unencapsulated drug removal step, e.g., by tangential flow filtration, dialysis, or size exclusion chromatography. Surprisingly, in an accelerated storage stability study of loaded liposome preparations that did not undergo buffer exchange, the phospholipids were more stable to degradation than in buffer-exchanged preparations. Analysis of ammonium in liposomes after loading with and without buffer exchange showed that buffer-exchanged liposomes indeed had significantly lower levels of ammonium in the extraliposomal medium. Therefore, the presence of ammonium in the extraliposomal medium ("exchanged ammonium") in an amount resulting from ammonium-drug transmembrane exchange during the drug loading step is preferable to reduce storage-related degradation of lipids. The inventors surprisingly found that even relatively small amounts, such as 1.7 mEq / L, of external (extraliposomal) ammonium have a significant stabilizing effect on liposomal lipids (Example 71). (For convenience, NH4 in mEq / L is used, where the salt form of ammonium is not defined.) +(The term "mM" is used to indicate concentration.) In some embodiments, the external medium of the liposome composition has ammonium or substituted ammonium less than 0.5 mEq / L. In some embodiments, the liposome composition contains ammonium or substituted ammonium in the extraliposome medium at a concentration of at least 1 mEq / L, at least 2 mEq / L, at least 5 mEq / L, at least 10 mEq / L, at least 15 mEq / L, or at least 20 mEq / L, but not more than 200 mEq / L, not more than 150 mEq / L, not more than 100 mEq / L, not more than 80 mEq / L, or not more than 60 mEq / L. In some embodiments, the liposome composition contains ammonium or substituted ammonium in the extraliposomal medium at a concentration of at least 1 mEq / L, at least 2 mEq / L, at least 5 mEq / L, at least 10 mEq / L, at least 15 mEq / L, or at least 20 mEq / L, and no more than 200 mEq / L, no more than 150 mEq / L, no more than 100 mEq / L, no more than 80 mEq / L, or no more than 60 mEq / L, and is stable against phospholipid degradation, with the extent of phospholipid degradation being less than 10%, less than 5%, or less than 1% of total phospholipids after 6 weeks at 37°C. In some embodiments, the liposome composition contains ammonium or substituted ammonium in the extraliposomal medium at a concentration of at least 1 mEq / L, at least 2 mEq / L, at least 5 mEq / L, at least 10 mEq / L, at least 15 mEq / L, or at least 20 mEq / L, and no more than 200 mEq / L, 150 mEq / L, 100 mEq / L, 80 mEq / L, or 60 mEq / L. The liposome composition is stable against phospholipid degradation, with the extent of phospholipid degradation being less than 10%, 7%, 5%, or 4% of total phospholipid after 3 months at 37°C. In some embodiments, the phospholipid is phosphatidylcholine. In some embodiments, the phospholipid is HSPC, and the ammonium salt is ammonium chloride, ammonium sulfate, or a combination thereof, at an ammonium concentration of 10-80 mM or 15-60 mM.In some embodiments, the phospholipid is HSPC and the ammonium salt is ammonium chloride, ammonium sulfate, or a combination thereof, at an ammonium concentration of 1-10 mM or 2-5 mM. In some embodiments, the normality of ammonium in the external medium of the liposome composition is within 90-110% of the normality of the encapsulated drug in the drug loading step, where normality is the concentration expressed in gram equivalents / L (eq / L).

[0144] The desired concentration of ammonium in the extraliposomal medium can be achieved by accumulation of extraliposomal ammonium during the drug loading step, at the expense of ammonium (used as part of the scavenger) that escapes from the liposomal interior, as explained above. Alternatively, the desired level of extraliposomal ammonium can be contributed by the extraliposomal ammonium remaining in the liposomes after removal of extraliposomal ammonium prior to drug loading, or by adding ammonium salts, such as ammonium chloride or ammonium sulfate, to the external medium of the liposomal formulation. In the latter case, if post-loading purification of the liposomes from unencapsulated material is desired, ammonium salts can be added to the liposomal preparation or to the exchange buffer after the post-loading buffer exchange / unencapsulated drug removal step.

[0145] In some embodiments, the liposome compositions provided herein may further comprise a lipophilic free radical scavenger, such as alpha-tocopherol, in the liposome formulation.

[0146] In some embodiments, the oxazolidinone liposome compositions provided herein comprise HSPC, cholesterol, and PEG-DSPE in a mass ratio of about 5:3:1. In some embodiments, the oxazolidinone liposome compositions provided herein comprise HSPC, cholesterol, and PEG-DSPE in a molar ratio of about 45:55:2.25. In some embodiments, the oxazolidinone liposome compositions comprise an oxazolidinone consisting of AKG-28 or a pharmaceutically acceptable salt thereof. In some embodiments, the oxazolidinone liposome compositions comprise an oxazolidinone consisting of AKG-38 or a pharmaceutically acceptable salt thereof.

[0147] [How to use] Disclosed herein are methods for inhibiting the growth of mycobacteria, such as Mycobacterium tuberculosis, or Gram-positive bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA). Additional mycobacteria and Gram-positive bacteria include, but are not limited to, Mycobacterium avium complex, Mycobacterium leprae, Mycobacterium gordonae, Mycobacterium abscessus, Mycobacterium mucogenicum, streptococci, vancomycin-resistant enterococci (VRE), Staphylococcus pneumoniae, Enterococcus faecium, Group B Streptococcus, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus viridans, Listeria monocytogenes, Nocardia, and Corynebacterium sp. In some embodiments, the compounds and compositions provided herein inhibit the growth of drug-resistant strains of Mycobacterium tuberculosis. In some embodiments, methods for treating mycobacterial infections are provided. In some embodiments, the compounds and compositions provided herein can be used to treat non-tuberculous mycobacterial infections. In some embodiments, the method comprises administering a therapeutically effective amount of an aminoalkyloxazolidinone and / or a pharmaceutically acceptable salt thereof of the present disclosure to a subject in need thereof. In some embodiments, the method comprises administering a therapeutically effective amount of a liposome composition comprising an aminoalkyloxazolidinone compound and / or a pharmaceutically acceptable salt thereof of the present disclosure to a subject in need thereof.

[0148] Mycobacteria are the genus of bacteria that cause tuberculosis (TB). According to the World Health Organization, TB is one of the top 10 causes of death worldwide and is the single leading cause of death from an infectious pathogen. Rifampicin is the most effective first-line drug for treating TB. However, there has been an increase in cases of infection with Mycobacterium tuberculosis strains that are resistant to rifampicin. Multidrug-resistant tuberculosis (MDR-TB) is a form of TB caused by bacteria that do not respond to isoniazid and rifampicin.

[0149] In some embodiments, the composition is a liquid pharmaceutical formulation for parenteral administration. In some embodiments, the liquid pharmaceutical formulation is a liposomal formulation containing an appropriate amount of the oxazolidinone compound described herein, wherein the oxazolidinone compound is encapsulated within the liposome. In another embodiment, the compound is in the form of a salt with a polyanion, such as sulfate, citrate, sucrose octasulfate, or inositol hexaphosphate, within the liposome. In some embodiments, the compound is a precipitated or gelled salt containing sulfate within liposomes composed of multiple lipid excipients, including, but not limited to, phosphatidylcholine, cholesterol, and PEGylated phosphatidylethanolamine. Liposomes according to the present disclosure exhibit entrapment efficiencies of greater than 85%, greater than 90%, and greater than 95%. In some embodiments, residual amounts of unentrapped drug are removed from the liposomal composition. This can be accomplished by various means, such as size exclusion chromatography, ion exchange, dialysis, ultrafiltration, tangential flow filtration, adsorption, or precipitation. During or after the unentrapped drug removal step, the liposomes can be incorporated into a desired pharmaceutically acceptable carrier, such as saline, isotonic dextrose, isotonic sucrose, Ringer's solution, or Hank's solution. A buffer substance can be added to achieve the desired physiologically acceptable pH. The liposome composition can be adjusted to the desired drug concentration and sterilized, for example, by sterile filtration through a 0.2-0.22 μm filter. In some embodiments, the compound concentration in the liposome composition ranges from 1-50 mg / ml, 3-30 mg / ml, or 5-25 mg / ml.

[0150] In some embodiments, pharmaceutical preparations containing the liposome compositions provided herein can be sterilized by conventional, well-known sterilization techniques. The aqueous solution can then be packaged for use or filtered under aseptic conditions and lyophilized, and the lyophilized preparation can be combined with a sterile aqueous solution before administration. The composition can contain pharmaceutically acceptable auxiliary substances necessary to approximate physiological conditions, such as pH adjusting and buffering agents, tonicity adjusting agents, etc., such as sodium acetate, sodium lactate, sodium chloride, potassium chloride, calcium chloride, etc. Furthermore, the lipid suspension can contain a lipid protective agent to protect lipids against damage caused by free radicals and lipid peroxides during storage. Lipophilic free radical quenchers such as alpha-tocopherol are suitable.

[0151] In some embodiments, the liposomes are mixed with one or more additional excipients for tonicity or pH control, including, but not limited to, sodium chloride, Hepes buffer, phosphate buffer, and histidine buffer.

[0152] The liposome composition may also contain other pharmaceutically acceptable substances required to approximate physiological conditions, such as pH adjusting and buffering agents, tonicity adjusting agents, etc., such as sodium acetate, sodium lactate, sodium chloride, potassium chloride, calcium chloride, etc. Furthermore, the liposome suspension may contain a lipid protective agent to protect lipids against damage caused by free radicals and lipid peroxides during storage. A lipophilic free radical quencher such as alpha-tocopherol is suitable.

[0153] In another embodiment, the composition is an oral formulation. In some embodiments, the composition is a liquid formulation. In some embodiments, the composition is a solid formulation (e.g., tablet, capsule, pill, dragee, caplet, etc.). For oral use, for example, tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, syrups, or elixirs can be prepared (Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.)). Compositions intended for oral use can be prepared according to any method known in the art for the manufacture of pharmaceutical compositions. The composition can contain one or more agents, such as antioxidants, sweeteners, flavoring agents, coloring agents, and preservatives, to provide a palatable preparation. Tablets containing the active ingredient mixed with non-toxic pharmaceutically acceptable excipients or adjuvants suitable for the manufacture of tablets are acceptable. Suitable excipients or adjuvants include, but are not limited to, for example, inert diluents, solubilizing agents, suspending agents, adjuvants, wetting agents, sweetening agents, flavoring or flavoring substances, isotonic substances, colloidal dispersing agents, and surfactants.

[0154] Tablets, dragees, capsules, pills, granules, suppositories, solutions, suspensions and emulsions, pastes, ointments, gels, creams, lotions, powders and sprays can be suitable pharmaceutical compositions.

[0155] The compounds or compositions may be administered topically, orally, parenterally, intraperitoneally, and / or rectally.

[0156] Dosage regimens are adjusted to provide the optimum desired response (e.g., a therapeutic response). For example, one or more doses can be administered over time, or the dose can be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation.

[0157] The dosage of the compounds and / or their pharmaceutically acceptable salts, or liposomes containing the compounds and / or their pharmaceutically acceptable salts, can vary within wide limits and will naturally have to be adjusted in each particular case to the individual conditions and the pathogens to be controlled.

[0158] In some embodiments, for use in treating bacterial infections, the compound or pharmaceutical liposomal composition is administered to a subject in need thereof once every 7 days (i.e., once every week), once every 14 days (i.e., once every 2 weeks), once every 21 days (i.e., once every 3 weeks), once every 28 days (i.e., once every 4 weeks), or once every 42 days (i.e., once every 6 weeks). In some embodiments, the average weekly dosage is from about 1 mg to about 1500 mg, from about 10 to about 700 mg, from about 25 to about 500 mg, or from about 70 to about 250 mg. In some embodiments, the average weekly dosage is about 1 mg to about 10 mg, about 10 mg to about 25 mg, about 25 mg to about 50 mg, about 50 mg to about 100 mg, about 100 mg to about 200 mg, about 200 mg to about 300 mg, about 300 mg to about 400 mg, about 400 mg to about 500 mg, about 500 mg to about 600 mg, about 600 mg to about 700 mg, about 700 mg to about 800 mg, about 800 mg to about 900 mg, about 900 mg to about 1000 mg, about 1000 mg to about 1100 mg, about 1100 mg to about 1200 mg, about 1200 mg to about 1300 mg, about 1300 mg to about 1400 mg, or about 1400 mg to about 1500 mg. In some embodiments, the compound or composition is administered for up to one month, up to two months, up to three months, up to four months, or more. The specific therapeutically effective amount depends on various factors, such as the bacterial infection being treated, the activity of the specific compound administered, the pharmaceutical composition employed, the age, weight, sex, etc., of the subject, the route of administration, the severity of the bacterial infection, optional drugs / active agents used in combination (sequentially or simultaneously) with the specific compound, and similar factors known to a practitioner of ordinary skill in the art. In some embodiments, the compound or composition can be used to treat tuberculosis or other mycobacterial infections. In some embodiments, the compound can be used as monotherapy. In some embodiments, treatment can include simultaneous and / or sequential administration of an effective amount of a compound described herein and an effective amount of one or more additional active agents for treating Mycobacterium tuberculosis and other gram-positive bacterial infections.In some embodiments, treatment can involve simultaneous and / or sequential administration of an effective amount of a compound described herein and two or more additional active agents (e.g., two, three, four, etc.) in effective amounts for treating Mycobacterium tuberculosis and other Gram-positive bacterial infections. A synergistic antibacterial effect refers to an antibacterial effect that is greater than the expected purely additive effect of the individual compounds of the combination. When administered simultaneously, the compound and the active agent can be contained in the same composition or in separate compositions. When administered sequentially, the composition containing the compound and the composition containing the additional active agent can be administered with a time separation (e.g., 20 minutes, 40 minutes, 60 minutes, or more). In some embodiments, the additional active agent can be administered using a different route of administration or by a different injection. For example, a compound according to the present disclosure can be administered intravenously, and one or more additional agents can be administered orally.

[0159] In some embodiments, administration of the compound with one or more (e.g., 1, 2, 3, or 4) additional active agents can result in a reduction in the length of the treatment period. For example, administration of the compound with one or more (e.g., 1, 2, 3, or 4) additional active agents can shorten the treatment period by at least 3 times, at least 2 times, or at least 1.5 times compared to treatment with only one active agent. In some embodiments, the additional agent(s) is an antibacterial agent. In some embodiments, the additional active agent includes, but is not limited to, fluoroquinolines such as moxifloxacin, gatifloxacin, or levofloxacin, bedaquiline and other diarylquinoline analogs (e.g., TBAJ-587 and TBAJ-876), delamanid, pretomanid, isoniazid, rifampicin, rifapentine, pyrazinamide, clofazimine, spectinamide, ethambutol, streptomycin, cannabinoids, fluticasone, fluoxetine, fluoxetine, fluoxetine-1, fluoxetine-2, fluoxetine-3, fluoxetine-4, fluoxetine-5, fluoxetine-6, fluoxetine-7, fluoxetine-8, fluoxetine-9, fluoxetine-10, fluoxetine-11, fluoxetine-12, fluoxetine-13, fluoxetine-14, fluoxetine-15, fluoxetine-15, fluoxetine-16, fluoxetine-17, fluoxetine-18, fluoxetine-19, fluoxetine-20, fluoxetine-21, fluoxetine-22, fluoxetine-23, fluoxetine-24, flu The inhibitors can include mycin, capreomycin, amikacin, the leucyl-tRNA synthetase (LeuRS) inhibitor GSK3036656, the tryptophan synthase inhibitor GSK839, the DprE1 inhibitors OPC-167832 and macodinone (PBTZ-169), Telacebec, GSK-656, TBA-7371, and amoxicillin plus clavulanate, pharmaceutically acceptable salts of each thereof, and any combination thereof. For the treatment of Gram-positive bacterial infections, the additional active agents may include, but are not limited to, vancomycin, gentamicin, daptomycin, teicoplanin, ceftaroline, ceftrobiprole, telavancin, dalbavancin, oritavancin, fluoroquinolines (e.g., delafloxacin), tetracyclines (e.g., eravacycline and omadacycline), sulfonamides (e.g., sulfamethoxazole), trimethoprim, lefamulin, and any combination thereof. In some embodiments, treatment may involve simultaneous and / or sequential administration of an effective amount of a compound described herein and an effective amount of bedaquiline, pretomanid, pyrazinamide, moxifloxacin, or a pharmaceutically acceptable salt of each thereof, or a combination of the foregoing.

[0160] Actual dosage levels of the active ingredients in the pharmaceutical compositions disclosed herein can be varied to provide an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration without causing toxicity to the patient.

[0161] "Parenteral," as used herein in the context of administration, means modes of administration other than enteral and topical administration, usually by injection, and includes, but is not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion.

[0162] "Parenteral administration" and "parenterally administered" as used herein refer to modes of administration other than enteral (i.e., via the digestive tract) and topical administration, usually by injection or infusion, and include, but are not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, inhalation, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion. Intravenous injection and infusion are often (but not exclusively) used for liposomal drug administration.

[0163] In some embodiments, the liquid composition is administered intravenously. In some embodiments, the compound or pharmaceutical composition is administered to a subject in need thereof once every 7 days (i.e., once every week), once every 14 days (i.e., once every 2 weeks), once every 21 days (i.e., once every 3 weeks), once every 28 days (i.e., once every 4 weeks), and once every 42 days (i.e., once every 6 weeks). In some embodiments, the average weekly dose is about 1 mg to about 1500 mg, about 10 to about 700 mg, about 25 to about 500 mg, or about 70 to about 250 mg. In some embodiments, the average weekly dose is about 1 mg to about 10 mg, about 10 mg to about 25 mg, about 25 mg to about 50 mg, about 50 mg to about 100 mg, about 100 mg to about 200 mg, about 200 mg to about 300 mg, about 300 mg to about 400 mg, about 400 mg to about 500 mg, about 500 mg to about 600 mg, about 600 mg to about 700 mg, about 700 mg to about 800 mg, about 800 mg to about 900 mg, about 900 mg to about 1000 mg, about 1000 mg to about 1100 mg, about 1100 mg to about 1200 mg, about 1200 mg to about 1300 mg, about 1300 mg to about 1400 mg, or about 1400 mg to about 1500 mg. The specific therapeutically effective amount will depend on a variety of factors, such as the bacterial infection being treated, the activity of the specific compound being administered, the pharmaceutical composition employed, the age, weight, sex, etc. of the subject, the route of administration, the severity of the bacterial infection, any optional drugs / active agents used in combination (sequentially or simultaneously) with the specific compound, and similar factors known to a physician of ordinary skill in the art.

[0164] In some embodiments, the liposome composition is administered parenterally.

[0165] In some embodiments, the method comprises simultaneously or sequentially administering one or more additional active agents, hi some embodiments, the one or more active agents comprise bedaquiline, pretomanide, pyrazinamide, moxifloxacin, pharmaceutically acceptable salts thereof, or combinations thereof.

[0166] In some embodiments, the liposome composition is administered once a week to once every six weeks.

[0167] In some embodiments, the percentage of the compound remaining in the blood after administration to a subject in need thereof is greater than 20% of the administered amount at 6 hours, hi some embodiments, the percentage of the compound remaining in the blood is greater than 10% of the administered amount.

[0168] One aspect of the present disclosure relates to a method of making a liposome composition, comprising the steps of: (i) preparing, in a medium substantially free of the entrapment agent, liposomes comprising phospholipids, cholesterol, and PEG-lipids, and having an interior space containing the entrapment agent; (ii) contacting the liposomes with a compound disclosed herein in an aqueous medium to effect encapsulation of the compound in the liposomes; (iii) removing unencapsulated compound; and (iv) providing the liposomes in a physiologically acceptable medium suitable for parenteral use.

[0169] In some embodiments, compound or pharmaceutical oral compositions is used for treating bacterial infection, administered once or twice a day.Specific therapeutically effective amount depends on various factors, for example, the bacterial infection to be treated, the activity of the specific compound to be administered, the pharmaceutical composition to be used, the age, weight, sex etc. of the subject, the route of administration, the severity of bacterial infection, the optional drug / active agent that is used in combination with specific compound (sequentially or simultaneously), and the similar factors that are known to the physician skilled in the art.

[0170] [Additional Embodiments] The following additional embodiments are provided for illustrative purposes. Lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 1.45:55:2.25 or a mass ratio of 5:3:1, and AKG-28 [ka] AKG-28 liposome composition comprising a pharmaceutically acceptable salt of 2. The composition of embodiment 1, wherein the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm. 3. The composition of embodiment 2, wherein the mono- or oligolamellar vesicles have a z-average diameter of 100 to 130 nm. 4. The composition of embodiment 1, wherein the liposome composition has a polydispersity index of less than 0.15. 5. The composition of embodiment 4, wherein the liposome composition has a polydispersity index of less than 0.10. 6. The composition of embodiment 1, wherein the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 230-290 g / mol. 7. The composition of embodiment 1, wherein the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 290-360 g / mol. 8. The composition of embodiment 1, wherein the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 300-340 g / mol. 9. The composition of embodiment 1, wherein the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is about 250 g / mol. 10. The composition of embodiment 1, wherein the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is about 330 g / mol. 11. The composition of embodiment 1, wherein the total concentration of AKG-28 in the composition is 8 to 15 mg / mL. 12. The composition of embodiment 1, wherein the total concentration of AKG-28 in the composition is 9 to 11 mg / mL. 13. The composition of embodiment 1, wherein the proportion of encapsulated AKG-28 relative to the total AKG-28 in the composition is at least 90%. 14. The composition of embodiment 1, wherein the proportion of encapsulated AKG-28 relative to the total AKG-28 in the composition is at least 95%. 15. The composition of embodiment 1, wherein the proportion of encapsulated AKG-28 relative to the total AKG-28 in the composition is at least 97%. 16. The composition of embodiment 1, wherein the proportion of encapsulated AKG-28 relative to the total AKG-28 in the composition is at least 98%. 17. The composition of embodiment 1, wherein the composition comprises liposome vesicles in an aqueous medium, the aqueous medium comprising sodium chloride and optionally ammonium ions. 18. The composition of embodiment 17, wherein the osmolality of the aqueous medium is 270 to 330 mOsmol / kg. 19. The composition of embodiment 17, wherein the osmolality of the aqueous medium is 270 to 310 mOsmol / kg. 20. The composition of embodiment 17, wherein the ammonium concentration in the aqueous medium is 20 to 60 mM. 21. The composition of embodiment 17, wherein the ammonium concentration in the aqueous medium is 50 to 80 mM. 22. The composition of embodiment 17, wherein the ammonium concentration in the aqueous medium is less than 0.5 mM. 23. The composition of embodiment 17, wherein the ammonium concentration in the aqueous medium is less than 130-150 mM. 24. The composition of embodiment 1, further comprising a buffer, wherein the buffer buffers the composition at a pH of 7.3 to 7.7. 25. The composition of embodiment 1, further comprising a buffer, wherein the buffer buffers the composition at a pH of about 7.5. 26. The composition of embodiment 1, further comprising a buffer, wherein the buffer buffers the composition at a pH of 7.5. 27. The composition of embodiment 1, further comprising HEPES or a phosphate buffer. 28. The composition of embodiment 27, wherein the composition comprises HEPES or phosphate buffer at a concentration of 5 to 50 mM. 29. The composition of embodiment 27, wherein the composition comprises HEPES or phosphate buffer at a concentration of about 20 mM. 30. The composition of embodiment 27, wherein the composition comprises HEPES or phosphate buffer at a concentration of 20 mM. 31. The composition of embodiment 1, further comprising a chelating agent. 32. The composition of embodiment 1, further comprising a chelating agent at a concentration of 0.1 to 1 mM. 33. The composition of embodiment 1, further comprising a chelating agent at a concentration of about 0.5 mM. 34. The composition of embodiment 1, further comprising a chelating agent at a concentration of 0.5 mM. 35. The composition of embodiment 1, further comprising deferoxamine (DFO) or EDTA. 36. The composition of embodiment 1, further comprising deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM. 37. The composition of embodiment 1, further comprising deferoxamine (DFO) or EDTA at a concentration of about 0.5 mM. 38. The composition of embodiment 1, further comprising deferoxamine (DFO) or EDTA at a concentration of 0.5 mM. 39. The composition of embodiment 1, wherein the composition is storage stable. 40. The composition of any one of embodiments 1-39, wherein the composition comprises AKG-28 as the sulfate salt of AKG-28. Lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 41.45:55:2.25 or a mass ratio of 5:3:1, and AKG-38 [ka] AKG-38 liposome composition comprising a pharmaceutically acceptable salt of 42. The composition of embodiment 41, wherein the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm. 43. The composition of embodiment 42, wherein the mono- or oligolamellar vesicles have a z-average diameter of 100 to 130 nm. 44. The composition of embodiment 41, wherein the liposome composition has a polydispersity index of less than 0.15. 45. The composition of embodiment 44, wherein the liposome composition has a polydispersity index of less than 0.10. 46. ​​The composition of embodiment 41, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 430-480 g / mol. 47. The composition of embodiment 41, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 500 to 650 g / mol. 48. The composition of embodiment 41, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 430 to 650 g / mol. 49. The composition of embodiment 41, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is about 450 g / mol. 50. The composition of embodiment 41, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is about 600 g / mol. 51. The composition of embodiment 41, wherein the total concentration of AKG-38 in the composition is 12 to 25 mg / mL. 52. The composition of embodiment 41, wherein the total concentration of AKG-38 in the composition is 13.5 to 16.5 mg / mL. 53. The composition of embodiment 41, wherein the total concentration of AKG-38 in the composition is about 15 mg / mL. 54. The composition of embodiment 41, wherein the total concentration of AKG-38 in the composition is about 20 mg / mL. 55. The composition of embodiment 41, wherein the proportion of encapsulated AKG-38 relative to the total AKG-38 in the composition is at least 90%. 56. The composition of embodiment 41, wherein the proportion of encapsulated AKG-38 relative to the total AKG-38 in the composition is at least 95%. 57. The composition of embodiment 41, wherein the proportion of encapsulated AKG-38 relative to the total AKG-38 in the composition is at least 97%. 58. The composition of embodiment 41, wherein the proportion of encapsulated AKG-38 relative to the total AKG-38 in the composition is at least 98%. 59. The composition of embodiment 41, wherein the composition comprises liposome vesicles in an aqueous medium, the aqueous medium comprising sodium chloride and optionally ammonium ions. 60. The composition of embodiment 59, wherein the osmolality of the aqueous medium is 270 to 330 mOsmol / kg. 61. The composition of embodiment 59, wherein the osmolality of the aqueous medium is 270 to 310 mOsmol / kg. 62. The composition of embodiment 59, wherein the ammonium concentration in the aqueous medium is 20 to 60 mM. 63. The composition of embodiment 59, wherein the ammonium concentration in the aqueous medium is 50 to 80 mM. 64. The composition of embodiment 59, wherein the ammonium concentration in the aqueous medium is less than 0.5 mM. 65. The composition of Example 59, wherein the ammonium concentration in the aqueous medium is less than about 0.5 mM. 66. The composition of embodiment 59, wherein the concentration of sodium chloride is 130-150 mM. 67. The composition of embodiment 41, further comprising a buffer, wherein the buffer buffers the composition at a pH of 7.3 to 7.7. 68. The composition of embodiment 41, further comprising a buffer, wherein the buffer buffers the composition at a pH of about 7.5. 69. The composition of embodiment 41, further comprising a buffer, wherein the buffer buffers the composition at a pH of 7.5. 70. The composition of embodiment 41, further comprising HEPES or a phosphate buffer. 71. The composition of embodiment 70, wherein the composition comprises HEPES or phosphate buffer at a concentration of 5 to 50 mM. 72. The composition of embodiment 70, wherein the composition comprises HEPES or phosphate buffer at a concentration of about 20 mM. 73. The composition of embodiment 70, wherein the composition comprises HEPES or phosphate buffer at a concentration of 20 mM. 74. The composition of embodiment 41, further comprising a chelating agent. 75. The composition of embodiment 41, further comprising a chelating agent at a concentration of 0.1 to 1 mM. 76. The composition of Example 41, further comprising a chelating agent at a concentration of about 0.5 mM. 77. The composition of embodiment 41, further comprising a chelating agent at a concentration of 0.5 mM. 78. The composition of embodiment 41, further comprising deferoxamine (DFO) or EDTA. 79. The composition of embodiment 41, further comprising deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM. 80. The composition of embodiment 41, further comprising deferoxamine (DFO) or EDTA at a concentration of about 0.5 mM. 81. The composition of embodiment 41, further comprising deferoxamine (DFO) or EDTA at a concentration of 0.5 mM. 82. The composition of embodiment 41, wherein the composition is storage stable. 83. The composition of any one of embodiments 1-82, comprising AKG-28 as the sulfate salt of AKG-28. Lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 84.45:55:2.25 or a mass ratio of 5:3:1, and AKG-28 [ka] AKG-28 liposome composition comprising a pharmaceutically acceptable salt of a. the liposome composition comprises mono- or oligolamellar vesicles having an average z-diameter of 90 to 130 nm; or the liposome composition comprises mono- or oligolamellar vesicles having an average z-diameter of 90 to 130 nm; or the liposome composition comprises mono- or oligolamellar vesicles having an average z-diameter of 100 to 130 nm; b. the liposome composition has a polydispersity index of less than 0.15; or the liposome composition has a polydispersity index of less than 0.10; c. The drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 230 to 280 g / mol; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 290 to 360 g / mol; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is 300 to 340 g / mol; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is about 250 g / mol; or the drug / lipid ratio of AKG-28 to total phospholipids (PhL) in the composition is about 330 g / mol; d. The total concentration of AKG-28 in the composition is 8 to 15 mg / mL; or the total concentration of AKG-28 in the composition is 9 to 11 mg / mL; e. The ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 90%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 95%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 97%; or the ratio of encapsulated AKG-28 to the total AKG-28 in the liposome composition is at least 98%; f. the liposome composition comprises an aqueous medium comprising sodium chloride and optionally comprising ammonium ions; g. The osmolality of the aqueous medium is 270-330 mOsmol / kg; or the osmolality of the aqueous medium is 270-310 mOsmol / kg; h. The ammonium concentration in the aqueous medium is 20 to 60 mM; or the ammonium concentration in the aqueous medium is 50 to 80 mM; or the ammonium concentration in the aqueous medium is less than 0.5 mM; or the ammonium concentration in the aqueous medium is less than 130 to 150 mM; i. the aqueous medium further comprises a buffer, and the buffer has a pH of 7.3 to 7.7; a pH of about 7.5; or buffers the liposome composition at a pH of 7.5; j. the aqueous medium further comprises HEPES or a phosphate buffer; or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 5 to 50 mM; or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of about 20 mM; or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 20 mM; k. The composition further comprises a chelating agent; or the composition further comprises a chelating agent at a concentration of 0.1-1 mM; or the composition further comprises a chelating agent at a concentration of about 0.5 mM; or the composition further comprises a chelating agent at a concentration of 0.5 mM; or the composition further comprises deferoxamine (DFO) or EDTA; or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1-1 mM; or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of about 0.5 mM; or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of 0.5 mM; or l.AKG-28 is encapsulated in liposomes as AKG-28 sulfate. The AKG-28 liposome composition is further characterized by any one or more of the following features: Lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 85.45:55:2.25 or a mass ratio of 5:3:1, and AKG-38 [ka] AKG-38 liposome composition comprising a pharmaceutically acceptable salt of a. the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90 to 130 nm; b. the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 100-130 nm; c. the liposome composition has a polydispersity index of less than 0.15, or the liposome composition has a polydispersity index of less than 0.10; d. The drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 430 to 480 g / mol, or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 500 to 650 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 430 to 650 g / mol; or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 450 g / mol, or the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the liposome composition is 600 g / mol; e. The total concentration of AKG-38 in the liposome composition is 12 to 25 mg / mL; or The total concentration of AKG-38 in the liposome composition is 13.5 to 16.5 mg / mL; or The total concentration of AKG-38 in the composition is about 15 mg / mL; or The total concentration of AKG-38 in the liposome composition is about 20 mg / mL; f. The ratio of encapsulated AKG-38 to the total AKG-38 in the liposome composition is at least 90%; or the ratio of encapsulated AKG-38 to the total AKG-38 in the liposome composition is at least 95%; or the ratio of encapsulated AKG-38 to the total AKG-38 in the liposome composition is at least 97%; or the ratio of encapsulated AKG-38 to the total AKG-38 in the liposome composition is at least 98%; g. The liposome composition comprises an aqueous medium comprising sodium chloride and optionally comprising ammonium ions; h. The osmolality of the aqueous medium is 270-330 mOsmol / kg, or the osmolality of the aqueous medium is 270-310 mOsmol / kg; i. The ammonium concentration in the aqueous medium is 20 to 60 mM; or the ammonium concentration in the aqueous medium is 50 to 80 mM; or the ammonium concentration in the aqueous medium is less than 0.5 mM; or the ammonium concentration in the aqueous medium is less than 0.5 mM; j. The concentration of sodium chloride is 130-150 mM; k. the aqueous medium further comprises a buffer, wherein the buffer buffers the liposome composition at a pH of 7.3 to 7.7, a pH of about 7.5, or a pH of 7.5; l. the aqueous medium further comprises HEPES or a phosphate buffer, or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 5 to 50 mM, or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of about 20 mM, or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 20 mM; m. the aqueous medium further comprises a chelating agent; or further comprises a chelating agent at a concentration of 0.1 to 1 mM, or further comprises a chelating agent at a concentration of about 0.5 mM, or further comprises a chelating agent at a concentration of 0.5 mM, or further comprises deferoxamine (DFO) or EDTA, or further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM, or further comprises deferoxamine (DFO) or EDTA at a concentration of about 0.5 mM, or further comprises deferoxamine (DFO) or EDTA at a concentration of 0.5 mM; n. The liposome composition is storage stable; or o.AKG-38 is encapsulated in liposomes as AKG-38 sulfate. The AKG-38 liposome composition is further characterized by any one or more of the following features: 86. AKG-28 liposomal dispersion formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof. 87. Liposomal dispersion formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one dihydrochloride monohydrate. 88. Isotonic liposomal dispersion of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one dihydrochloride monohydrate. 89. The liposomal dispersion according to any one of embodiments 86 to 88, comprising unilamellar lipid bilayer liposomal vesicles encapsulating an aqueous space containing (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one as the sulfate salt. 90. The liposomal dispersion of embodiment 89, wherein the liposomal vesicles are composed of hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and (PEG(molecular weight 2,000)-distearoylphosphatidylethanolamine, PEG-DSPE) (PEG(2000)-DSPE). 91. The liposome dispersion according to embodiment 90, comprising 45 to 65 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 92. The liposome dispersion according to embodiment 90, comprising 50 to 60 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 93. The liposome dispersion according to embodiment 90, comprising about 50 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 94. The liposome dispersion according to embodiment 90, comprising about 55 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 95. The liposomal dispersion according to embodiment 90, wherein the lipid content of the liposomal dispersion consists of HSPC, and PEG(2000)-DSPE. 96. The liposomal dispersion of embodiment 95, comprising HSPCs and cholesterol in a weight ratio of about 5:3. 97. The liposomal dispersion according to embodiment 95, comprising HSPC and cholesterol in a molar ratio of about 45:55. 98. The liposomal dispersion of embodiment 95, comprising HSPC, cholesterol and PEG(2000)-DSPE in a weight ratio of about 5:3:1. 99. The liposome dispersion of embodiment 95, comprising HSPC, cholesterol and PEG(2000)-DSPE in a molar ratio of about 45:55:2.25. 100. The liposomal dispersion of any one of embodiments 87 to 99, further comprising 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES) buffer or phosphate buffer. 101. The liposomal dispersion of embodiment 100, further comprising sodium chloride at a concentration of 50 to 80 mM. 102. The liposomal dispersion of any one of embodiments 99-101, further comprising ammonium substituted during the drug loading process at a concentration of 20-60 mM. 103. The liposome dispersion according to any one of embodiments 99 to 102, wherein the liposome vesicles have a z-average diameter of 90 to 130 nm. 104. AKG-38 liposomal dispersion formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof. 105. Liposomal dispersion formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one hydrochloride. 106. Isotonic liposomal dispersion of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one hydrochloride. 107. A liposomal dispersion according to any one of embodiments 104 to 106, comprising unilamellar lipid bilayer vesicles encapsulating an aqueous space containing (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one as the sulfate salt. 108. The liposome dispersion described in embodiment 107, wherein the liposome vesicles are composed of hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and (PEG(molecular weight 2,000)-distearoylphosphatidylethanolamine, PEG-DSPE) (PEG(2000)-DSPE). 109. The liposome dispersion according to embodiment 108, comprising 50 to 65 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 110. The liposome dispersion according to embodiment 108, comprising 50 to 60 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 111. The liposome dispersion according to embodiment 108, comprising about 50 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 112. The liposome dispersion according to embodiment 108, comprising about 55 mol% cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 113. The liposomal dispersion according to embodiment 108, wherein the lipid content of the liposomal dispersion consists of HSPC and PEG(2000)-DSPE. 114. The liposomal dispersion according to embodiment 113, comprising HSPCs and cholesterol in a weight ratio of about 5:3. 115. The liposomal dispersion according to embodiment 113, comprising HSPCs and cholesterol in a molar ratio of about 45:55. 116. The liposome dispersion according to embodiment 113, comprising HSPC, cholesterol and PEG(2000)-DSPE in a weight ratio of about 5:3:1. 117. The liposome dispersion according to embodiment 113, comprising HSPC, cholesterol and PEG(2000)-DSPE in a molar ratio of about 45:55:2.25. 118. The liposomal dispersion according to any one of embodiments 104 to 117, further comprising 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES) buffer or phosphate buffer. 119. The liposomal dispersion of embodiment 118, further comprising sodium chloride at a concentration of 50 to 80 mM. 120. The liposomal dispersion of any one of embodiments 118-119, further comprising ammonium substituted during the drug loading process at a concentration of 20-60 mM. 121. The liposome dispersion according to any one of embodiments 118 to 120, wherein the liposomes have a z-average diameter of 90 to 130 nm. 122. A liposomal dispersion according to any one of embodiments 86 to 120, wherein the dispersion comprises liposomal vesicles having a z-average diameter of 90 to 130 nm. 123. A liposomal dispersion according to any one of embodiments 86 to 121, wherein the dispersion comprises liposomal vesicles having a z-average diameter of 100 to 130 nm. 124. A liposomal dispersion according to any one of embodiments 86 to 121, wherein the dispersion comprises liposomal vesicles having a z-average diameter of about 100 nm. 125. The liposomal dispersion of any one of embodiments 86-124, wherein the dispersion further comprises a chelating agent. 126. The liposomal dispersion of any one of embodiments 86-124, wherein the dispersion further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA. 127. The liposomal dispersion of any one of embodiments 86 to 124, wherein the dispersion further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA at a concentration of 0.1 to 1 mM. 128. The liposomal dispersion of any one of embodiments 86-124, wherein the dispersion further comprises a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA at a concentration of 0.5 mM. 129. The liposomal dispersion according to any one of embodiments 86 to 128, wherein the dispersion further comprises ammonium at a concentration of 20 to 60 mM. 130. The liposomal dispersion according to any one of embodiments 86 to 129, wherein the dispersion further comprises sodium chloride at a concentration of 50 to 80 mM. 131. An AKG-28 liposome dispersion having a pH of 7.0 to 8.0, wherein the liposome dispersion is a. unilamellar lipid bilayer vesicles containing phospholipids and greater than 50 mol% cholesterol; and b. (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one as the sulfate salt formed within liposome vesicles AKG-28 liposomal dispersion comprising: 132. The liposomal dispersion of embodiment 131, formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one hydrochloride. 133. The liposome dispersion according to embodiment 132, comprising 45 to 65 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 134. The liposome dispersion according to embodiment 132, comprising 50 to 55 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 135. The liposome dispersion according to embodiment 132, comprising about 50 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 136. The liposome dispersion according to embodiment 132, comprising about 55 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 137. The liposome dispersion according to embodiment 132, wherein the liposome dispersion is obtained by a process comprising the steps of dissolving (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one hydrochloride in a drug loading solution, and contacting the drug loading solution with extracted purified liposome vesicles containing an ammonium sulfate sequestering agent to load AKG-28 into the liposome vesicles. 138. The liposome dispersion according to any one of embodiments 131 to 137, wherein the unilamellar lipid bilayer vesicles comprise HSPCs and cholesterol in a molar ratio of 45:55 or a mass ratio of 5:3. 139. The liposome dispersion according to embodiment 138, wherein the unilamellar lipid bilayer vesicles consist of HSPC, cholesterol, and PEG(2000)-DSPE. 140. The liposome dispersion according to embodiment 139, wherein the unilamellar lipid bilayer vesicles consist of HSPC, cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1. 141. The liposomal dispersion according to any one of embodiments 131 to 140, having a pH of 141.7 to 7.7. 142. An AKG-28 liposome dispersion having a pH of 7.0 to 8.0, wherein the liposome dispersion is a. Unilamellar lipid bilayer vesicles containing phospholipids and at least 45 mol% cholesterol; b. (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one as the sulfate salt formed within liposome vesicles; and c. a chelating agent selected from the group consisting of deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM AKG-28 liposomal dispersion comprising: 143. An AKG-38 liposome dispersion having a pH of 7.0 to 8.0, wherein the liposome dispersion is a. unilamellar lipid bilayer liposome vesicles comprising phospholipids and at least 50 mol% cholesterol; and b. (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one as the sulfate salt formed within liposome vesicles AKG-38 liposomal dispersion comprising: 144. The liposomal dispersion of embodiment 143, formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one hydrochloride. 145. The liposome dispersion according to any one of embodiments 143 to 144, wherein the liposome dispersion is obtained by a process comprising the steps of dissolving (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one hydrochloride in a drug loading solution, and contacting the drug loading solution with extracted purified liposome vesicles containing an ammonium sulfate sequestering agent to load AKG-38 into the liposome vesicles. 146. The liposome dispersion according to any one of embodiments 144 to 145, wherein the lipid vesicles comprise HSPC and cholesterol in a molar ratio of 45:55 or a mass ratio of 5:3. 147. The liposome dispersion according to embodiment 146, wherein the lipid vesicles consist of HSPC, cholesterol and PEG(2000)-DSPE. 148. The liposome dispersion according to embodiment 147, wherein the lipid vesicles consist of HSPC, cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1. 149. The liposomal dispersion according to any one of embodiments 146 to 148, having a pH of 149.7 to 7.7. 150. An AKG-38 liposome dispersion having a pH of 7-8, wherein the liposome dispersion is a. Unilamellar lipid bilayer vesicles containing phospholipids and at least 50 mol% cholesterol; b. (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one as the sulfate salt formed within liposome vesicles; and c. a chelating agent selected from the group consisting of deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM AKG-38 liposomal dispersion comprising: 151. The liposomal dispersion of embodiment 150, formulated with the hydrochloride salt of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one. 152. The liposome dispersion according to embodiment 151, wherein the liposome dispersion is obtained by a process comprising the steps of dissolving the hydrochloride salt of (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one in a drug loading solution, and contacting the drug loading solution with extracted purified liposomes containing an ammonium sulfate sequestering agent to load AKG-38 into liposome vesicles. 153. The liposome dispersion according to any one of embodiments 150 to 152, wherein the lipid unilamellar lipid bilayer vesicles comprise HSPCs and cholesterol in a molar ratio of 45:55 or a mass ratio of 5:3. 154. The liposome dispersion according to embodiment 153, wherein the unilamellar lipid bilayer vesicles consist of HSPC, cholesterol, and PEG(2000)-DSPE. 155. The liposome dispersion according to embodiment 154, wherein the unilamellar lipid bilayer vesicles consist of HSPC, cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1. 156. The liposome dispersion according to any one of embodiments 150 to 155, having a pH of 156.7 to 7.7. 157. The liposome dispersion according to any one of embodiments 142 to 145 and 150 to 152, comprising 50 to 65 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 158. The liposome dispersion according to any one of embodiments 142 to 145 and 150 to 152, comprising 50 to 55 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome vesicles. 159. The liposome dispersion according to any one of embodiments 142-145 and 150-152, comprising about 50 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. 160. The liposome dispersion according to any one of embodiments 142-145 and 150-152, comprising about 55 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipids in the liposome vesicles. An oxazolidinone liposome dispersion having a pH of 161.7 to 8, wherein the liposome dispersion is a. Unilamellar lipid bilayer vesicles containing phospholipids and more than 50 mol% cholesterol; b. Formula (I) as a sulfate salt formed within liposome vesicles [ka] [In the formula, R1 is a tetrazole ring substituted at the 2' position with an aminoalkyl; R2 is an amine (NH2) or acetamide (NHCOCH3)] oxazolidinone; and c. a chelating agent selected from the group consisting of deferoxamine (DFO) or EDTA at a concentration of 0.1 to 1 mM 1. An oxazolidinone liposomal dispersion comprising: 162. The liposomal dispersion according to embodiment 161, comprising phospholipid and cholesterol in a mass ratio of about 5 to 3. 163. The liposomal dispersion according to embodiment 161, further comprising PEG-DSPE. 164. The liposome dispersion according to embodiment 163, wherein the liposome composition comprises phospholipid, cholesterol and PEG-DSPE in a mass ratio of about 5:3:2.25. 165. The liposome dispersion according to embodiment 164, wherein the liposome composition comprises phospholipid, cholesterol and PEG-DSPE in a molar ratio of about 45:55:2.25 mol%. 166. The liposomal dispersion according to any one of embodiments 161-165, wherein the phospholipid is HSPC. 167. The liposomal dispersion according to any one of embodiments 161 to 166, wherein the oxazolidinone is (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof. 168. The liposomal dispersion according to any one of embodiments 161 to 166, wherein the oxazolidinone is (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof. 169. The liposomal dispersion according to any one of embodiments 167-168, wherein the oxazolidinone is a sulfate salt formed within liposomal vesicles that comprise an ammonium sulfate (AS) sequestering agent within the liposomal dispersion. 170. The liposome dispersion according to any one of embodiments 161 to 169, wherein the unilamellar lipid bilayer vesicles comprise a total of 55 mol% cholesterol relative to the total amount of phospholipids in the liposome dispersion. 171. The liposomal dispersion according to any one of embodiments 161 to 170, wherein the phospholipids are present in a total of 45 mol % cholesterol relative to the total amount of phospholipids in the liposomal dispersion. 172. The liposomal dispersion according to any one of embodiments 161-171, wherein the phospholipids are present in a total of 45 mol % cholesterol relative to the total amount of phospholipids in the liposomal dispersion. 173. The liposomal dispersion according to any one of embodiments 161-172, wherein the oxazolidinone is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof: [Table 7] JPEG2025501769000034.jpg245160JPEG2025501769000035.jpg255160JPEG2025501769000036.jpg199160174. A liposome dispersion comprising lipid vesicles formed from (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof, and phospholipids, 55 mol% cholesterol and 5 mol% PEG-DSG. 175. A liposome dispersion having a pH of 7 to 8, comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof, and lipid vesicles formed from phospholipids and 55 mol% cholesterol. 175. A liposomal dispersion comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylamino)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof; lipid vesicles formed from phospholipids and 55 mol% cholesterol; and a chelating agent. 176. The liposomal dispersion of embodiment 175, wherein the chelating agent is selected from the group consisting of deferoxamine (desferrioxamine, desferal), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). 177. A liposome dispersion comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof, and lipid vesicles formed from phospholipids, 55 mol% cholesterol, and 5 mol% PEG-DSG. 178. A liposome dispersion having a pH of 7 to 8, comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof, and lipid vesicles formed from phospholipids and 55 mol% cholesterol. 179. A liposomal dispersion comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof; lipid vesicles formed from phospholipids and 55 mol% cholesterol; and a chelating agent. 180. The liposomal dispersion of embodiment 179, wherein the chelating agent is selected from the group consisting of deferoxamine (desferrioxamine, desferal), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). 181. A liposome dispersion comprising (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharmaceutically acceptable salt thereof, and lipid vesicles formed from phospholipids, 55 mol% cholesterol, and 5 mol% PEG-DSG.

[0171] [Additional Embodiments] Further embodiments The following additional embodiments are provided for illustrative purposes and include the oxazolidinone liposome compositions described in the additional embodiments below, and other combinations of the features recited therein. having a pH of 1.7.3 to 7.7; a. a chelating agent selected from the group consisting of DFO, EDTA, and DTPA; b. liposome vesicles comprising phospholipids and greater than 40 mol% cholesterol relative to the total phospholipids in the liposome composition; and c. Formula (I) encapsulated in liposome vesicles [ka] [In the formula, R2 is an amine (NH2) or acetamide (NHCOCH3), R1 is a tetrazole ring substituted at the 2' position with an aminoalkyl. A liposomal pharmaceutical composition comprising the sulfate salt of the compound of formula (I). 2. The liposome composition of embodiment 1, wherein the aminoalkyl is dimethylaminoethyl. 3. The liposome vesicle has the formula 1b: [ka] 2. The liposome composition of embodiment 1, comprising the compound: 4. The liposome vesicle is a compound of formula 1c [ka] 2. The liposome composition of embodiment 1, comprising the compound: 5. The liposome vesicle is represented by Formula 1d or Formula 1e [ka] 2. The liposome composition of embodiment 1, comprising a compound of formula: 6. The liposome composition according to any one of embodiments 1 to 5, wherein the liposome vesicles are in an aqueous medium. 7. The liposome composition of any one of embodiments 1 to 5, wherein the compound is entrapped in the liposome vesicle along with an entrapment agent, and the entrapment agent comprises a polyanion. 8. The liposome composition of embodiment 7, wherein the sequestering agent is sucrose triethylammonium octasulfate or ammonium sulfate. 9. The liposome composition of embodiment 7, wherein the sequestering agent is sucrose triethylammonium octasulfate. 10. The liposome composition of embodiment 7, wherein the sequestering agent is ammonium sulfate. 11. The liposome composition of any one of embodiments 1 to 5, comprising a salt of the compound, wherein the salt is a sulfate, a citrate, a sucrosophate, a salt with a phosphorylated or sulfated polyol, or a salt with a phosphorylated or sulfated polyanionic polymer. 12. The liposome composition of any one of embodiments 1 to 5, comprising a salt of the compound, wherein the salt is a sulfate salt. 13. The liposome composition according to any one of embodiments 1 to 5, wherein the compound in the liposome vesicle has an aqueous solubility of less than 1 mg / mL. 14. The composition of any one of embodiments 1 to 5, wherein the compound in the liposome vesicle has an aqueous solubility of less than 0.1 mg / mL. 15. The liposome composition of any one of embodiments 1 to 5, wherein the liposome vesicles comprise a membrane comprising phosphatidylcholine and cholesterol. 16. The liposome composition according to any one of embodiments 1 to 5, wherein a membrane separates the interior of the liposome vesicle from the aqueous medium. 17. The liposome composition of embodiment 15, wherein the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soy phosphatidylcholine (HSPC). 18. The liposome composition of embodiment 15, wherein the molar ratio of phosphatidylcholine to cholesterol is from about 60:40 to about 35:65. 19. The liposome composition of embodiment 15, wherein the molar ratio of phosphatidylcholine to cholesterol is from about 55:45 to about 35:65. 20. The liposome composition of embodiment 15, wherein the molar ratio of phosphatidylcholine to cholesterol is from about 50:50 to about 45:55. 21. The liposome composition of embodiment 15, wherein the molar ratio of phosphatidylcholine to cholesterol is from about 50:50 to about 40:60. 22. The liposome composition according to any one of embodiments 15 to 21, wherein the membrane further comprises a polymer-conjugated lipid. 23. The liposome composition of any one of embodiments 1 to 5, wherein the liposome vesicles comprise HSPC, cholesterol, and polymer-conjugated lipid in a molar ratio of about 45:55:2.75. 24. The liposome composition of embodiment 22 or embodiment 23, wherein the polymer-conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE). 25. The liposome composition according to any one of embodiments 1 to 5, wherein the composition is a liquid pharmaceutical formulation for parenteral administration. 26. The liposome composition of any one of embodiments 1 to 5, wherein the liposomes have a Z-average particle size in the range of about 80 to about 130 nm. 27. A method for treating a bacterial infection, comprising administering to a subject in need thereof a therapeutically effective amount of the liposome composition of any one of embodiments 1-5. 28. The method of embodiment 27, wherein the bacterial infection is a Mycobacterium tuberculosis infection. 29. The method of embodiment 27 or embodiment 28, wherein the compound in the liposome vesicle has a minimum inhibitory concentration (MIC) in the range of about 0.01 μg / ml to about 0.25 μg / ml. 30. The method of embodiment 27 or embodiment 28, wherein the compound in the liposome vesicle has an MIC in the range of about 0.01 μg / ml to about 0.1 μg / ml. 31. The method of any one of embodiments 27-30, comprising parenterally administering the liposome composition. 32. The method of embodiment 31, wherein the method comprises administering one or more active agents simultaneously or sequentially. 33. The method of embodiment 32, wherein the one or more active agents comprises bedaquiline, pretomanid, pyrazinamide, moxifloxacin, pharmaceutically acceptable salts thereof, or combinations thereof. 34. The method of embodiment 31, wherein the liposome composition is administered once a week to once every six weeks. 35. The method of embodiment 31, wherein the percentage of the compound remaining in the blood after administration to a subject in need thereof is greater than 20% of the administered amount in 6 hours. 36. The method of embodiment 31, wherein the percentage of the compound remaining in the blood after administration to a subject in need thereof is greater than 10% of the administered amount. 37. A method of making a liposome composition, comprising: (i) preparing liposomes comprising phospholipids, cholesterol, and PEG-lipids in a medium substantially free of the entrapment agent, the liposomes having an interior space containing the entrapment agent; (ii) contacting the liposome with a compound according to any one of embodiments 1 to 8 in an aqueous medium to encapsulate the compound in the liposome; (iii) removing any unencapsulated compound; (iv) providing the liposomes in a physiologically acceptable medium suitable for parenteral use; A method comprising: [Example]

[0172] The following examples, including experiments conducted and results achieved, are provided for illustrative purposes only and are not to be construed as limiting the present disclosure.

[0173] [Example 1] [Synthesis of oxazolidinone derivatives] Compounds AKG-1, AKG-2, AKG-6, AKG-8, AKG-9, and AKG-19 were synthesized by reacting tedizolid mesylate (tedizolid-MS) with the respective amines in N-methyl-2-pyrrolidone (NMP) as a solvent at 60 °C (Scheme 1). Tedizolid-MS was obtained by mesylation of the 1° hydroxyl group of tedizolid with methanesulfonyl chloride in the presence of a base at room temperature (RT). Treatment of tedizolid-MS with sodium azide followed by reduction of the resulting azide (AKG-3-A) afforded intermediate 1 as the free base or AKG-3 as the hydrochloride salt, depending on the eluent selected for purification. Amidation of intermediate 1 with the corresponding acid followed by hydrochloride salt formation using HCl / EtOAc afforded compounds AKG-17 and AKG-18. Reaction of tedizolid with the corresponding dialkylamino acids under standard esterification conditions gave compounds AKG-5 and AKG-20. O-Alkylation of tedizolid with 2-chloro-N,N-diethylaminoethylamine using sodium hydride as the base gave compound AKG-7.

[0174] Intermediate 2 was synthesized by boronation of commercially available aryl bromides using bis(pinocolato)diboron (Scheme 2). Suzuki coupling of intermediate 2 with readily available 5-bromo-2-fluoropyridine gave intermediate 3, which was heated with the corresponding amine in NMP in a sealed tube to give compounds AKG-11–AKG-15.

[0175] Compounds AKG-16, AKG-21 to AKG-27 were prepared in a convergent synthesis starting from intermediate 4 (Schemes 3 and 4). Click chemistry using sodium azide on 5-bromo-2-cyanopyridine gave intermediate 4. N-Alkylation of tetrazole in intermediate 4 gave intermediates 5 and 6 in a 3:1 ratio. The structures of these intermediates were deduced from HMBC analysis. Intermediates 7 to 12 were synthesized, and regioisomers were obtained in a similar manner (only the desired isomer is shown in Scheme 4). Suzuki coupling of intermediates 5 to 12 with intermediate 2 and deprotection of the amine group, where applicable, gave compounds AKG-16, AKG-21 to AKG-27.

[0176] Intermediate 13 was synthesized by mesylation of readily available aryl bromides. Intermediate 14 was reduced with hydrazine to give intermediate 15 (Scheme 5). Boc protection or acetylation of the primary amine in intermediate 15, followed by boronation, gave intermediates 18 and 19, respectively. Suzuki coupling of the boronic acid intermediate with the corresponding aryl bromide intermediate (U.S. Patent Application Publication No. 2010 / 0022772, WO 2013 / 044845, incorporated herein by reference in their entireties) and deprotection of the amine group, if applicable, gave compounds AKG-28 to AKG-31 and AKG-38 to AKG-40.

[0177] [Synthetic scheme] For the synthesis of intermediate 19, see U.S. Patent Application Publication No. 2010 / 0022772, WO 2013 / 044845, which are incorporated by reference in their entireties.

[0178] FIG. 24 shows the synthesis scheme 1.

[0179] FIG. 25 shows the synthesis scheme 2.

[0180] FIG. 26 shows the synthesis scheme 3.

[0181] FIG. 27 shows the synthesis scheme 4.

[0182] Figure 28 shows the synthesis scheme 5.

[0183] [Synthesis] (material and method) Tedizolid, (R)-3-(4-bromo-3-fluorophenyl)-5-(hydroxymethyl)oxazolidin-2-one, was purchased from Skychemical, dimethyl-(2-piperidin-4-yl-ethyl)-amine was purchased from Enamine, and other reagents and solvents were purchased from Adams and used as received. The chemical structure of the final product was confirmed by nuclear magnetic resonance spectroscopy (NMR) determined on a Bruker NMR spectrometer (500 MHz or 400 MHz). 1 H NMR, 13 It was characterized by C NMR. 13 C NMR spectra were fully resolved, and chemical shifts were expressed in parts per million (ppm) using the deuterated solvent peak or tetramethylsilane (internal) as the internal standard. 1 H NMR data are reported as follows: chemical shift (d, ppm), multiplicity (s, singlet; br s, broad singlet; d, doublet; t, triplet; m, multiplet), integral, coupling constant (Hz). 13 C NMR data are reported in units of chemical shift (d, ppm). The purity of the final product (≥95%) was confirmed by analytical HPLC. Analytical HPLC was performed on an Agilent analytical HPLC system using a Sunfire column, 3.5 μm (150 cm × 4.6 mm), a gradient system (water (0.01% TFA) / ACN (0.01% TFA)) and a flow rate of 1 mL / min, with detection at 254 and 214 nm. Flash chromatography (FC) purification was performed using silica gel 60 (0.04–0.063 nm; 230–400 mesh) from Santai Technologies.

[0184] Procedure A. A reaction mixture of tedizolid-MS (1.0 equiv.), R1R2NH (4.0 equiv.) in NMP (10 mL) was heated to 60° C. in a sealed tube for 15 h. Upon completion (LCMS), the reaction was diluted with HO (40 mL) and extracted with EtOAc (2×50 mL). The combined extracts were washed with saturated brine, dried over Na2SO4, and filtered. The solvent was removed under reduced pressure, and the residue was purified using FC to give the product in greater than 95% purity.

[0185] 1. Synthesis of Tedizolid-Ms

[0186] [ka] To a solution of tedizolid (7.00 g, 18.90 mmol) and triethylamine (3.83 g, 37.80 mmol) in CHCl (50 mL) at 0° C. under Ar was added methanesulfonyl chloride (3.25 g, 28.36 mmol) dropwise at 0° C. After stirring at RT for 2 h, the reaction mixture was poured into water and extracted with CHCl. ​​The organic layer was washed with brine, dried over NaSO, and collected by filtration. The solvent was removed under reduced pressure to give the pure product tedizolid-Ms (7.0 g, 82.6% yield) as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.31 - 8.14 (m, 2H), 7.88 - 7.65 (m, 2H), 7.53 (d, J= 8.6 Hz, 1H), 5.14 - 4.96 (m, 1H), 4.59 - 4.39 (m, 5H), 4.28 (t, J = 9.4 Hz, 1H), 3.92 (dd, J = 9.2, 6.3 Hz, 1H), 3.28 (s, 3H). MS (ESI+) m / z 449.1 ([M + 1] + ).

[0187] (2. Synthesis of AKG-1, 2, 6, 8, 9 and 19)

[0188] [ka] Using procedure A, AKG-1 was obtained from tedizolid-Ms and dimethylamine as a white solid (0.5 g, 56.4% yield). 1 H NMR (400 MHz, DMSO-d6) δ 8.94 (s, 1H), 8.32 - 8.13 (m, 2H), 7.83 - 7.64 (m, 2H), 7.54 (d, J= 7.6 Hz, 1H), 4.87 (s, 1H), 4.49 (s, 3H), 4.21 (t, J = 8.6 Hz, 1H), 3.84 (t, J= 7.4 Hz, 1H), 2.62 (s, 2H), 2.25 (s, 6H). 13 C NMR (101 MHz, DMSO-d6) δ164.3, 161.0, 158.6, 154.6, 149.9, 145.5, 140.9, 137.6, 132.1, 131.4, 122.6, 119.1, 114.6, 106.0, 72.0, 62.1, 48.7, 46.4, 40.2. MS (ESI+) m / z 398.2 ([M + 1] + ).

[0189] [ka]

[0190] Using procedure A, AKG-2 was obtained from tedizolid-Ms and dimethylamine as a white solid (0.52 g, 54.8% yield). 1 H NMR (400 MHz, DMSO-d6) δ 8.94 (s, 1H), 8.29 - 8.11 (m, 2H), 7.81 - 7.65 (m, 2H), 7.52 (dd, J = 8.6, 1.8 Hz, 1H), 4.89 - 4.73 (m, 1H), 4.49 (s, 3H), 4.19 (t, J = 8.8 Hz, 1H), 3.82 (dd, J = 8.7, 7.0 Hz, 1H), 2.75 (dd, J = 5.1, 3.7 Hz, 2H), 2.57 (q, J = 6.9 Hz, 4H), 0.97 (t, J = 7.1 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.3, 161.0, 158.6, 154.7, 149.9, 145.5, 141.0, 137.6, 132.1, 131.3, 122.5, 119.1, 114.6, 106.1, 72.6, 56.1, 48.6, 47.7, 40.3, 12.3. MS (ESI+) m / z 426.3 ([M + 1] + ).

[0191] [ka]

[0192] Using procedure A, AKG-6 was obtained from tedizolid-Ms and N,N-dimethyl-2-(piperidin-4-yl)ethan-1-amine as a white solid (0.66 g, 58.2% yield). 1H NMR (400 MHz, CDCl3) δ 8.93 (s, 1H), 8.30 (dd, J = 8.1, 2.4 Hz, 1H), 8.05 (d, J = 7.8 Hz, 1H), 7.62 (d, J = 12.9 Hz, 1H), 7.56 - 7.47 (m, 1H), 7.45 - 7.37 (m, 1H), 4.89 - 4.74 (m, 1H), 4.48 (s, 3H), 4.11 (t, J = 8.6 Hz, 1H), 3.86 (t, J = 7.8 Hz, 1H), 2.93 (dd, J = 28.8, 10.9Hz, 2H), 2.80 - 2.64 (m, 2H), 2.50 - 2.04 (m, 11H), 1.69 (d, J= 10.8 Hz, 2H), 1.48 (d, J = 7.1 Hz, 2H), 1.37 - 1.19 (m, 4H). 13 C NMR (101 MHz, CDCl3) δ 164.7 , 161.3, 158.8 , 154.3 , 149.9 , 145.4 , 140.2 , 137.0, 132.3 , 130.5 , 122.0 , 120.0 , 113.8 , 106.4, 71.5, 61.4, 57.0, 55.3, 54.3, 48.9, 45.0, 39.7, 33.6, 32.4. MS (ESI+) m / z 509.2 ([M + 1] + ).

[0193] [ka]

[0194] Tedizolid-Ms and N using procedure A 1 ,N 1 -Diethylpropane-1,3-diamine gave AKG-8 as a white solid (0.62 g, 57.6% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.94 (s, 1H), 8.34 - 8.11 (m, 2H), 7.84 - 7.59 (m, 2H), 7.52 (dd, J = 8.6, 2.0 Hz, 1H), 4.80 (dd, J= 8.3, 5.7 Hz, 1H), 4.49 (s, 3H), 4.18 (t, J= 8.9 Hz, 1H), 3.90 (dd, J = 8.8, 6.5 Hz, 1H), 2.94 - 2.77 (m, 2H), 2.66 - 2.53 (m, 7H), 1.65 - 1.51 (m, 2H), 0.99 (t, J = 7.1 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.3, 161.0, 158.6, 154.6, 149.9, 145.5, 141.0, 137.6, 132.1, 131.4, 122.6, 119.1, 114.6, 106.1, 73.2, 52.1, 50.7, 48.2, 48.0, 46.7, 40.3, 26.4, 11.4. m / z 483.2 ([M + 1] + ).

[0195] [ka]

[0196] Tedizolid-Ms and N using procedure A 1 ,N 1 -diethylethane-1,2-diamine to give AKG-9 as a white solid (0.36 g, 34.4% yield). 1H NMR (500 MHz, DMSO-d6) δ 8.94 (s, 1H), 8.26 - 8.16 (m, 2H), 7.78 - 7.66 (m, 2H), 7.53 (d, J= 8.5 Hz, 1H), 4.85 - 4.73 (m, 1H), 4.49 (s, 3H), 4.18 (t, J = 8.8 Hz, 1H), 3.90 (t, J= 7.5 Hz, 1H), 2.88 (t, J = 5.4 Hz, 2H), 2.65 (t, J = 6.1 Hz, 2H), 0.95 (t, J = 7.0 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.3, 161.0, 158.6, 154.7, 149.9, 145.5, 141.0, 137.6, 132.1, 131.4, 122.6, 119.1, 114.6, 106.1, 73.3, 52.6, 52.2, 48.1, 47.6, 47.1, 40.3, 12.0. m / z 469.3 ([M + 1] + ).

[0197] Using procedure A, AKG-19 was obtained from tedizolid-Ms and ethane-1,2-diamine as a white solid (0.60 g, 55% yield). 1 H NMR (500 MHz, DMSO-d6) δ 10.33 (s, 1H), 9.89 (s, 1H), 8.95 (s, 1H), 8.58 (s, 3H), 8.23 ​​(q, J= 8.3 Hz, 2H), 7.79 (t, J = 8.8 Hz, 1H), 7.69 (d, J = 13.5 Hz, 1H), 7.49 (d, J = 8.7 Hz, 1H), 5.25 - 5.19 (m, 1H), 4.49 (s, 3H), 4.33 (t, J = 9.2 Hz, 1H), 4.05 (dd, J = 9.1, 6.7 Hz, 1H), 3.52 (s, 2H), 3.43 - 3.23 (m, 4H). 13C NMR (101 MHz, DMSO-d6) δ164.26, 160.94, 158.50, 153.79, 149.84, 145.51, 140.58, 137.78, 132.04, 131.42, 122.61, 119.50, 114.94, 106.46, 69.38, 49.59, 47.87, 45.16, 40.34, 35.58.

[0198] (3. Synthesis of AKG-3)

[0199] [ka] To a solution of tedizolid-Ms (1.00 g, 2.23 mmol) in DMF (20 mL) was added NaN (0.44 g, 6.69 mmol). After stirring at 90 °C for 3 h, the reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, dried over anhydrous MgSO, filtered, and concentrated under reduced pressure. The residue was further purified by column chromatography to give the title compound AKG-3-1 (0.7 g, 79.4% yield) as a white solid.

[0200] [ka]

[0201] A reaction mixture of AKG-3-1 (0.7 g, 1.77 mmol) and PhP (1.39 g, 5.31 mmol) in HO (2 mL) and THF (20 mL) was heated to reflux for 1 h. After completion (LCMS), the reaction was concentrated under reduced pressure and purified using reverse-phase FC. Purification using 0-10% MeOH in DCM as the eluent and lyophilization afforded the free base Intermediate 1 (2.5 g, 76.5% yield) as a yellow solid. FC purification using 0-30% MeCN in 0.006 M HCl in HO as the eluent afforded the hydrochloride salt AKG-3 (0.35 g, 48.8% yield) as a yellow solid after lyophilization. 1H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.61 (s, 3H), 8.28 - 8.18 (m, 2H), 7.79 (t, J= 8.8 Hz, 1H), 7.69 (dd, J = 13.5, 2.1 Hz, 1H), 7.48 (dd, J = 8.6, 2.1 Hz, 1H), 5.13 - 5.00 (m, 1H), 4.49 (s, 3H), 4.29 (t, J = 9.2 Hz, 1H), 4.02 (dd, J= 9.3, 6.6 Hz, 1H), 3.34 - 3.23 (m, 2H). 13 C NMR (101 MHz, DMSO-d6) δ163.8, 160.4, 158.0, 153.4, 149.4, 145.1, 140.2, 137.2, 131.5, 130.9, 122.1, 118.9, 114.3, 105.9, 69.8, 47.1, 41.4, 39.8. m / z 370.3 ([M -HCl + 1] + ).

[0202] (4.AKG-17 synthesis)

[0203]

change

[0204] (5. Synthesis of AKG-18)

[0205] [ka] Using the procedure for AKG-17, AKG-18 was obtained from intermediate 1 and 4-((tert-butoxycarbonyl)amino)butanoic acid as a yellow solid (0.5 g, 37.6% yield). 1 H NMR (500 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.52 (t, J = 5.7 Hz, 1H), 8.29 - 8.08 (m, 5H), 7.77 (t, J = 8.8 Hz, 1H), 7.69 (d, J = 13.6 Hz, 1H), 7.50 (d, J = 8.7 Hz, 1H), 4.87 - 4.76 (m, 1H), 4.50 (s, 3H), 4.22 (t, J = 9.0 Hz, 1H), 3.93 - 3.83 (m, 1H), 3.49 (t, J= 5.3 Hz, 2H), 2.83 - 2.72 (m, 2H), 2.28 (t, J = 7.2 Hz, 2H), 1.88 - 1.75 (m, 2H). 13 C NMR (101 MHz, DMSO-d6) δ 172.59, 164.23, 160.96, 158.52, 154.44, 149.00, 145.39, 140.88, 137.78, 132.10, 131.40, 122.61, 119.17, 114.70, 106.21, 72.17, 47.78, 41.88, 40.37, 38.78, 32.44, 23.60.

[0206] (6. Synthesis of AKG-5)

[0207] [ka] To a mixture of tedizolid (1.0 g, 2.70 mmol), 4-(dimethylamino)butanoic acid hydrogen chloride (0.57 g, 3.37 mmol), and TEA (0.27 g, 2.70 mmol) under N was added a catalytic amount of DMAP, DCC (0.84 g, 4.05 mmol) in DMF (20 mL). The mixture was stirred at RT for 16 h. Upon completion of the reaction (LCMS), it was diluted with HO (100 mL) and filtered. The filtrate was acidified to pH = 5-6 with 0.02 M HCl and then purified using RP-FC (eluent: MeCN in 0.5% formic acid / HO) to give the product AKG-5 as the formate salt after lyophilization. The product was redissolved in HO, and 1 equivalent of aqueous HCl (0.02 M) was added. The product was lyophilized to give AKG-5 as the HCl salt (600 mg, 42.7% yield). 1 H NMR (400 MHz, DMSO-d6) δ10.40 (br, 1H), 8.95 (s, 1H), 8.23 ​​(q, J= 8.5 Hz, 2H), 7.78 (t, J = 8.8 Hz, 1H), 7.71 (dd, J = 13.6, 2.1 Hz, 1H), 7.53 (dd, J = 8.6, 2.1 Hz, 1H), 5.03 (dd, J = 5.6, 3.1 Hz, 1H), 4.48 (s, 3H), 4.36 (qd, J = 12.4, 4.2 Hz, 2H), 4.26 (t, J = 9.3 Hz, 1H), 3.95 (dd, J = 9.2, 6.2 Hz, 1H), 2.99 - 2.86 (m, 2H), 2.64 (s, 6H), 2.45 (t, J = 7.3 Hz, 2H), 1.87 (m, 2H). 13C NMR (126 MHz, DMSO-d6) δ 172.3, 164.3, 158.8, 154.3, 149.9, 145.6, 140.8, 137.7, 132.0, 131.5, 122.6, 119.4, 114.7, 106.2, 71.1, 64.8, 56.3, 46.7, 40.3, 30.9, 19.9. m / z 469.3 ([M + 1] + ). m / z 484.1 ([M -HCl + 1] + ).

[0208] (7. Synthesis of AKG-7)

[0209] [ka] To a mixture of tedizolid (1.0 g, 2.70 mmol) in DMF (20 mL) was added NaH (0.13 g, 60%, 5.40 mmol) at RT under N. The mixture was stirred at 0 °C for 0.5 h, after which 2-diethylaminoethyl chloride hydrochloride (930 mg, 5.40 mmol) was added in one portion. The entire mixture was stirred at RT for 3 h. LCMS showed the reaction was complete. The reaction was carefully poured into ice / HO (20 mL) and extracted with DCM (2 × 50 mL). The combined organic extracts were washed with saturated brine and then dried over NaSO. The solvent was removed under reduced pressure, and the residue was purified using FC (eluent with MeOH in DCM 0-15%) to give AKG-7 as a white solid (0.5 g, 39.4% yield). 11H NMR (500 MHz, CDCl3) δ 8.93 (s, 1H), 8.30 (d, J = 8.2 Hz, 1H), 8.05 (d, J = 8.2 Hz, 1H), 7.72 (d, J = 12.9 Hz, 1H), 7.53 (t, J = 8.5 Hz, 1H), 7.42 (d, J = 8.5 Hz, 1H), 4.88 (d, J = 3.5 Hz, 1H), 4.48 (s, 3H), 4.34 - 4.26 (m, 1H), 4.18 - 4.08 (m, 2H), 4.00 - 3.93 (m, 1H), 3.87 (qd, J = 10.8, 2.9 Hz, 2H), 3.19 - 3.11 (m, 2H), 3.06 (q, J = 7.1 Hz, 4H), 1.26 (t, J = 7.2 Hz, 6H). 13 13C NMR (126 MHz, CDCl3) δ 164.7, 161.1, 159.1, 154.3, 149.8, 145.5, 140.0, 137.0, 132.2, 130.6, 122.0, 120.1, 113.8, 106.3, 71.3, 71.3, 66.9, 51.9, 48.2, 46.%, 39.7, 8.9. m / z 470.3([M + 1] + ).

[0210] (Synthesis of 8.AKG-20)

[0211]

Chemical Structure

[0210] (Synthesis of 8.AKG-20)

[0211]

Chemical Structure

[0212] 9. Synthesis of Intermediate 3

[0213] [ka] A mixture of (R)-3-(4-bromo-3-fluorophenyl)-5-(hydroxymethyl)oxazolidin-2-one (9.0 g, 31.02 mmol), bis(pinacolato)diboron (11.88 g, 46.54 mmol), and KOAc (4.56 g, 46.54 mmol) in dioxane (200 mL) was purged with Ar for 10 minutes, and then (PhP)PdCl (1.09 g, 1.55 mmol) was added. The mixture was purged with Ar again, and then heated to 90 °C for 15 hours. LCMS showed the reaction was complete. It was cooled to room temperature and filtered over Celite to give intermediate 2 as the filtrate. To the filtrate was added 5-bromo-2-fluoropyridine (6.55 g, 37.22 mmol), K3PO4 (14.47 g, 6.80 mmol), and HO (20 mL). The mixture was purged with Ar for 10 minutes, and (dppf)PdCl2 (2.27 g, 3.10 mmol) was added. The mixture was purged with Ar again. It was then heated to 90 °C for 15 hours. The reaction was monitored by LCMS. Upon completion, it was concentrated under reduced pressure, and the residue was diluted with HO (200 mL) and extracted with EtOAc (2 x 200 mL). The combined extracts were washed with saturated brine and then dried over Na2SO4. Filtration and removal of the solvent under reduced pressure gave a residue which was purified using FC (eluent with MeOH in DCM 0-15%) to give the product intermediate 3 (6.8 g, 71.6% yield for two steps) as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.43 (s, 1H), 8.23 ​​- 8.14 (m, 1H), 7.72 - 7.61 (m, 2H), 7.49 (dd, J = 8.6, 2.2 Hz, 1H), 7.32 (dd, J= 8.6, 2.7 Hz, 1H), 5.27 (t, J = 5.6 Hz, 1H), 4.80 - 4.71 (m, 1H), 4.15 (t, J= 9.1 Hz, 1H), 3.90 (dd, J = 8.9, 6.1 Hz, 1H), 3.75 - 3.67 (m, 1H), 3.63 - 3.55 (m, 1H).MS (ESI+) m / z 307 ([M + 1]+ ).

[0214] (10. Synthesis of AKG-11, 12, 13, 14, and 15) Procedure B. Intermediate 3 (1.0 equiv.), R1R2NH (4.0 equiv.), and a catalytic amount of DMAP in NMP (10 mL) were heated to 100 °C in a sealed tube for 16 h. Upon completion of the reaction (LCMS), it was diluted with HO (50 mL) and extracted with EtOAc (2 x 50 mL). The combined organic extracts were washed with saturated brine, dried over Na2SO4, and filtered. The solvent was removed under reduced pressure, and the residue was purified using RPFC (0.1% NH4HCO3 / MeCN in HO, 0-40%, eluent with C18) to give the product.

[0215] [ka]

[0216] Using procedure B, AKG-11 was obtained from intermediate 3 and N,N-dimethyl-2-(piperidin-4-yl)ethan-1-amine as a white solid (0.40 g, 30.1% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.28 (s, 1H), 7.73 - 7.65 (m, 1H), 7.60 (dd, J = 13.6, 2.1 Hz, 1H), 7.54 (t, J = 8.9 Hz, 1H), 7.41 (dd, J = 8.6, 2.1 Hz, 1H), 6.89 (d, J = 9.0 Hz, 1H), 5.25 (t, J = 5.6 Hz, 1H), 4.78 - 4.68 (m, 1H), 4.33 (d, J = 13.0 Hz, 2H), 4.12 (t, J = 9.0 Hz, 1H), 3.87 (dd, J = 8.9, 6.2 Hz, 1H), 3.74 - 3.64 (m, 1H), 3.62 - 3.52 (m, 1H), 2.87 - 2.71 (m, 2H), 2.23 (t, J = 7.3 Hz, 2H), 2.11 (s, 6H), 1.72 (d, J = 11.5 13C NMR (101 MHz, DMSO-d6) δ 160.68, 158.33, 154.81, 147.54, 139.15, 137.82, 130.32, 120.72, 119.11, 114.35, 106.97, 106.03, 105.74, 73.82, 62.09, 56.98, 46.45, 45.73, 45.39, 34.29, 34.15, 31.94.MS (ESI+) m / z 443.1 ([M + 1] + ).

[0217] [ka]

[0218] Use procedure B. Intermediate 3 and N 1 ,N 1 -dimethylethane-1,2-diamine to give AKG-12 as a white solid (0.52 g, 42.6% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.16 (s, 1H), 7.64 - 7.46 (m, 3H), 7.39 (dd, J = 8.6, 2.2 Hz, 1H), 6.58 (dd, J = 9.9, 5.6 Hz, 2H), 5.25 (t, J = 5.6 Hz, 1H), 4.80 - 4.66 (m, 1H), 4.11 (t, J = 9.0 Hz, 1H), 3.86 (dd, J = 8.9, 6.2 Hz, 1H), 3.76 - 3.65 (m, 1H), 3.61 - 3.49 (m, 1H), 3.37 (dd, J = 12.3, 6.5 Hz, 2H), 2.42 (t, J= 6.6 Hz, 2H), 2.18 (s, 6H). 13 C NMR (101 MHz, DMSO-d6) δ160.60, 158.48, 158.19, 154.82, 147.57, 138.92, 137.10, 130.22, 121.18, 118.53, 114.30, 108.37, 106.02, 105.74, 73.81, 62.10, 58.76, 46.45, 45.76, 39.21. MS (ESI+) m / z 375.1 ([M + 1] + ).

[0219] [ka]

[0220] Use procedure B. Intermediate 3 and N 1 ,N 1 -Diethylethane-1,2-diamine gave AKG-13 as a white solid (0.68 g, 51.9% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.17 (s, 1H), 7.67 - 7.46 (m, 3H), 7.39 (dd, J = 8.6, 2.1 Hz, 1H), 6.63 - 6.44 (m, 2H), 5.25 (s, 1H), 4.74 (dd, J = 9.2, 5.8 Hz, 1H), 4.12 (t, J= 9.0 Hz, 1H), 3.87 (dd, J = 8.9, 6.2 Hz, 1H), 3.76 - 3.65 (m, 1H), 3.63 - 3.52 (m, 1H), 3.34 (dd, J = 13.2, 6.2Hz, 2H), 2.60 - 2.55 (m, 2H), 2.54 - 2.50 (m, 4H), 0.97 (t, J= 7.1 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 160.60, 158.53, 158.18, 154.81, 147.62, 138.91, 137.13, 130.20, 121.16, 118.54, 114.29, 108.24, 105.87, 73.80, 62.10, 52.19, 47.13, 46.45, 39.48, 12.31. MS (ESI+) m / z 417.1 ([M + 1] + ).

[0221] [ka]

[0222] Use procedure B. Intermediate 3 and N 1 ,N 1 -dimethylpropane-1,3-diamine to give AKG-14 as a white solid (0.6 g, 47.3% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.16 (s, 1H), 7.63 - 7.54 (m, 2H), 7.50 (t, J = 8.9 Hz, 1H), 7.39 (dd, J = 8.6, 2.2 Hz, 1H), 6.73 (t, J = 5.6 Hz, 1H), 6.54 (d, J = 8.7 Hz, 1H), 5.25 (t, J = 5.5 Hz, 1H), 4.78 - 4.68 (m, 1H), 4.11 (t, J = 9.0 Hz, 1H), 3.87 (dd, J = 8.9, 6.2 Hz, 1H), 3.75 - 3.66 (m, 1H), 3.64 - 3.53 (m, 1H), 3.33 - 3.23 (m, 2H), 2.28 (t, J = 7.1 Hz, 2H), 2.13 (s, 6H), 1.72 - 1.62 (m, 2H). 13 C NMR (101 MHz, DMSO-d6) δ 160.60, 158.62, 158.18, 154.81, 147.62, 138.89, 137.08, 130.19, 121.21, 118.39, 114.29, 108.10, 106.02, 105.74, 73.81, 62.10, 57.44, 46.45, 45.72, 39.58, 27.54. MS (ESI+) m / z 389.1 ([M + 1] + ).

[0223] [ka]

[0224] Use procedure B. Intermediate 3 and N 1 ,N 1 -Diethylpropane-1,3-diamine gave AKG-15 as a white solid (0.65 g, 48.0% yield). 11H NMR (400 MHz, DMSO-d6) δ 8.16 (s, 1H), 7.63 - 7.54 (m, 2H), 7.50 (t, J = 8.9 Hz, 1H), 7.39 (dd, J = 8.6, 2.2 Hz, 1H), 6.75 (t, J = 5.5 Hz, 1H), 6.54 (d, J = 8.7 Hz, 1H), 5.25 (t, J = 5.4 Hz, 1H), 4.79 - 4.68 (m, 1H), 4.12 (t, J = 9.0 Hz, 1H), 3.87 (dd, J = 8.9, 6.2 Hz, 1H), 3.75 - 3.66 (m, 1H), 3.63 - 3.54 (m, 1H), 3.32 - 3.23 (m, 2H), 2.49 - 2.40 (m, 6H), 1.70 - 1.61 (m, 2H), 0.95 (t, J = 7.1 Hz, 6H). 13 13C NMR (101 MHz, DMSO-d6) δ 160.60, 158.65, 158.18, 154.81, 147.64, 138.89, 137.05, 130.18, 121.21, 118.37, 114.29, 108.01, 106.02, 105.74, 73.80, 62.09, 50.81, 46.80, 46.45, 40.11, 27.10, 12.23. MS (ESI+) m / z 417.1 ([M + 1] + ).

[0225] (Synthesis of 11.AKG-16)

[0226]

Chemical Structure

[0227] [ka]

[0228] A mixture of intermediate 4 (10.0 g, 44.25 mmol) and Ca(OH) (7.20 g, 97.35 mmol) in HO (150 mL) and DMF (20 mL) was stirred at rt for 0.5 h, after which (2-bromoethyl)dimethylamine hydrobromide (25.0 g, 107.3 mmol) was added. The mixture was heated at 80 °C for 24 h. LCMS showed a 3:1 mixture of intermediates 5 and 6, respectively. The mixture was diluted with HO (40 mL) and extracted with EtOAc (2 × 50 mL). The combined extracts were washed with saturated brine, dried over NaSO, and filtered. The solvent was removed under reduced pressure, and the residue was purified using FC (eluent with MeOH in DCM 0-15%) to give the crude product. The crude product was further purified by RPFC (MeCN 0-30% in 0.1% NH4HCO3 / HO, C18, intermediate 5 eluted first, then intermediate 6) to give intermediate 5 (0.74 g, 5.6% yield) as a white solid and intermediate 6 (0.25 g, as a light yellow solid).

[0229] Intermediate 5: 1 H NMR (400 MHz, DMSO-d6) δ 8.89 (dd, J = 2.3, 0.6 Hz, 1H), 8.27 (dd, J = 8.4, 2.4 Hz, 1H), 8.10 (dd, J = 8.4, 0.6 Hz, 1H), 4.87 (t, J = 6.1 Hz, 2H), 2.87 (t, J = 6.1 Hz, 2H), 2.17 (s, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 163.74, 151.48, 145.51, 140.81, 124.40, 122.21, 57.73, 51.54, 45.29. MS (ESI+) m / z 297.1, 299.1 ([M + 1] + ). Intermediate 6: 1H NMR (400 MHz, DMSO-d6) δ 8.98 (s, 1H), 8.38 (dd, J = 8.4, 2 Hz, 1H), 8.20 (d, J = 8.4 Hz, 1H), 5.00 (t, J = 6.4 Hz, 2H), 2.75 (t, J = 6 Hz, 2H), 2.10 (s, 6H), MS (ESI+) m / z 297.1, 299.1 ([M + 1] + ).

[0230] [ka]

[0231] A mixture of freshly prepared Intermediate 2 (1.68 g, 4.98 mmol) (from 1.44 g of (R)-3-(4-bromo-3-fluorophenyl)-5-(hydroxymethyl)oxazolidin-2-one using the procedure for Intermediate 3), Intermediate 5 (740 mg, 2.49 mmol), and KPO (1.16 g, 5.48 mmol) in dioxane (50 mL) and HO (5 mL) was purged with Ar for 10 min. To this was added (dppf)PdCl (182 mg, 0.25 mmol). The mixture was purged with Ar again. It was then heated to 90 °C for 15 h. LCMS showed the reaction was complete. It was concentrated under reduced pressure, and the residue was diluted with HO (200 mL) and extracted with EtOAc (2 × 200 mL). The combined extracts were washed with saturated brine, dried over NaSO, and filtered. The solvent was removed under reduced pressure, and the residue was purified using RPFC (eluent with MeCN in HO, 0-40%) to give the product AKG-16 (520 mg, 49.0% yield) as a white solid. 1H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.23 ​​(dd, J = 18.3, 8.2 Hz, 2H), 7.82 - 7.66 (m, 2H), 7.54 (dd, J = 8.6, 2.1 Hz, 1H), 5.28 (s, 1H), 4.89 (t, J = 6.1 Hz, 2H), 4.82 - 4.71 (m, 1H), 4.17 (t, J= 9.0 Hz, 1H), 3.98 - 3.87 (m, 1H), 3.77 - 3.65 (m, 1H), 3.64 - 3.53 (m, 1H), 2.90 (t, J = 6.1 Hz, 2H), 2.19 (s, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.17, 161.02, 158.58, 154.81, 149.91, 145.59, 141.03, 137.63, 132.10, 131.39, 122.59, 119.05, 114.47, 105.97, 105.69, 73.95, 62.07, 57.72, 51.46, 46.46, 45.26. MS (ESI+) m / z 428.1 ([M + 1] + ).

[0232] (12.AKG-21 synthesis)

[0233]

change

[0234] (13. Synthesis of AKG-22)

[0235] [ka] To a mixture of intermediate 7 (500 mg, 1.354 mmol) in HO (2 mL) and dioxane (8 mL) was added intermediate 2 (685 mg, 2.03 mmol), KPO (862 mg, 4.06 mmol), and (dppf)PdCl (99 mg, 0.135 mmol). The flask was evacuated and backfilled with Ar. The mixture was then stirred at 90 °C for 16 h. Water (20 mL) was added and extracted with EtOAc (2 × 20 mL). The organic layer was washed with brine, dried over NaSO, filtered, and concentrated. The residue was purified by silica gel column chromatography (Biotage, 40 g silica gel column @ 30 mL / min, eluted with 0–100% EtOAc in petroleum ether) to give the desired product AKG-22-1 (450 mg, yield: 66%) as a gray solid.

[0236] [ka]

[0237] To a mixture of AKG-22-1 (450 mg, 0.9 mmol) in DCM (8 mL) was added 4 M HCl / dioxane (2 mL). The mixture was then stirred at RT for 5 h. The solvent was removed under reduced pressure to give the desired product AKG-22 (390 mg, yield: 99%) as a gray solid. 1 H NMR (400 MHz, DMSO-d6) δ 9.03 (s, 1H), 8.39 (d, J = 8.0 Hz, 1H), 8.32 (d, J = 8.0 Hz, 1H), 8.19 (brs, 3H), 7.82 - 7.70 (m, 2H), 7.57 (dd, J = 8.8, 2.0 Hz, 1H), 5.18 (t, J = 5.8 Hz, 2H), 4.81 - 4.74 (m, 1H), 4.17 (t, J = 9.2 Hz, 1H), 3.93 (dd, J = 9.2, 6.4 Hz, 1H), 3.71 (dd, J = 12.4, 3.2 Hz, 1H), 3.59 (dd, J = 12.4, 4.0 Hz, 1H), 3.53 - 3.47 (m, 2H). 13 C NMR (400MHz, DMSO-d6) δ (161.07,158.63), 154.82, 152.52, 149.54, 143.25, (141.39,141.28), 138.24, 124.54, (118.66,118.53), 114.60, (106.01,105.73), 73.97, 62.02, 47.37, 46.48, 38.84.

[0238] (14. Synthesis of AKG-23)

[0239] [ka] To intermediate 8 (1.0 g, 2.71 mmol) in 20 mL of 1,4-dioxane and 5 mL of HO was added intermediate 2 (4.86 mmol, 1.63 g), KPO (1.14 g, 5.42 mmol), and (dppf)PdCl (0.23 g, 0.27 mmol), and the mixture was stirred at 100 °C for 16 h. After the starting material was consumed, 100 mL of saturated NaHCO was added. The aqueous phase was extracted with EtOAc (3 × 30 mL), and the combined organic extracts were washed with HO, concentrated under reduced pressure, and purified by FC to give the desired compound AKG-23-1 (1.0 g, 70% yield).

[0240] [ka]

[0241] To AKG-23-1 (1.0 g, 2 mmol) in 30 mL of DCM was added 1 mL of HCl (4 M in 1,4-dioxane), and the mixture was stirred for 1 hour. After the starting material was consumed, the mixture was filtered to obtain the crude product. The crude product was stirred in 3 mL of MeOH for 1 hour and filtered to obtain the desired product AKG-23 (0.53 g, 63% yield) as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.97 (s, 1H), 8.26 (m, 5H), 7.83-7.64 (m, 2H), 7.54 (dd, J = 8.6, 1.9 Hz, 1H), 5.09 (s, 2H), 4.77 (m, 1H), 4.16 (t, J = 9.1 Hz, 1H), 3.92 (dd, J= 8.8, 6.2 Hz, 1H), 3.71 (dd, J = 12.3, 3.2 Hz, 1H), 3.61-3.57 (dd, J = 12.3, 3.2 Hz, 1H), 3.53 (m, 3H). 13C NMR (125 MHz, DMSO-d6) δ38.31, 46.49, 50.86, 62.01, 73.96, [105.69, 105.97], 114.46, [118.90, 119.03], 122.73, [131.37, 131.47], 132.21, [137.66, 137.70], [140.99, 141.10], 145.42, 149.86, 154.82, [158.57, 161.02], 164.50.

[0242] (15. Synthesis of AKG-24)

[0243] [ka] A mixture of intermediate 2 (1.66 g, 4.98 mmol), intermediate 9 (800 mg, 2.49 mmol), and KPO (1.16 g, 5.48 mmol) in dioxane (50 mL) and HO (5 mL) was purged with Ar for 10 minutes, and (dppf)PdCl (182 mg, 0.25 mmol) was added. The mixture was purged with Ar again. It was then heated to 90 °C for 15 hours. LCMS showed the reaction was complete; it was concentrated under reduced pressure, and the residue was diluted with HO (200 mL) and extracted with EtOAc (2 × 200 mL). The combined extracts were washed with saturated brine, then dried over NaSO, and filtered. The solvent was removed under reduced pressure, and the residue was purified using RPFC (eluent with MeCN in HO, 0-40%) to give the product AKG-24 (440 mg, 39.0% yield) as a white solid. 1H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.28 - 8.16 (m,2H), 7.81 - 7.67 (m,2H), 7.54 (dd, J = 8.6, 2.1 Hz, 1H), 5.27 (t, J = 5.6 Hz, 1H), 4.93 - 4.70 (m, 3H), 4.17 (t, J = 9.1 Hz, 1H), 3.92 (dd, J = 8.9, 6.1 Hz, 1H), 3.72 (m, 1H), 3.60 (m, 1H), 3.04 (s, 2H), 0.87 (t, J = 6.9 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.13, 161.03, 158.59, 154.81, 149.94, 149.90, 145.66, 141.09, 140.98, 137.66, 137.62, 132.10, 132.08, 131.41, 131.37, 122.54, 119.15, 119.02, 114.50, 114.47, 105.99, 105.71, 73.95, 62.08, 52.09, 51.60, 46.85, 46.48, 12.26.MS (ESI+) m / z 456 ([M + H] + ).

[0244] (16.AKG-25 synthesis)

[0245]

change

[0246] [ka]

[0247] A mixture of intermediate 10 (0.74 g, 2.4 mmol), intermediate 2 (1.62 g, 4.8 mmol), and KPO (1 g, 4.8 mmol) in dioxane (30 mL) and HO (5 mL) was purged with Ar for 10 min, and Pd(dppf)Cl (175 mg, 0.24 mmol) was added. The mixture was purged with Ar again and heated to 90 °C for 15 h. It was concentrated under reduced pressure, and the residue was diluted with HO (80 mL) and extracted with EtOAc (2 × 100 mL). The combined extracts were washed with saturated brine, then dried over NaSO, and filtered. The solvent was removed under reduced pressure, and the residue was purified using FC (eluent with MeOH in DCM 0–15%) to give the product AKG-25 (0.73 g, 69.5% yield) as a gray solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.94 (s, 1H), 8.24-8.26 (m, 1H), 8.19-8.21 (m, 1H), 7.78-7.70 (m, 2 H), 7.54 (dd, J = 8.4, 2.0 Hz, 1H), 5.27 (t, J=5.6 Hz, 1H), 4.80 (t, J = 6.8 Hz, 2H), 4.77-4.75 (m, 1H), 4.16 (t, J = 9.2 Hz, 1H), 3.92 (dd, J=8.8 Hz, 6.0 Hz, 1H), 3.74 - 3.69 (m, 1H), 3.63 - 3.58 (m, 1H), 2.28 (t, J = 7.2 Hz, 2H), 2.10 - 2.17 (m, 8H). 13 C NMR (101 MHz, DMSO-d6) δ 164.26, 161.03, 158.58, 154.81, 149.92, 145.58, 141.09, 137.65, 132.10, 131.37, 122.60, 119.12, 118.99, 114.46, 105.98, 105.70, 73.95, 62.07, 55.96, 51.65, 46.47, 45.53, 27.21. + ).

[0248] (17. Synthesis of AKG-26)

[0249] [ka] To a solution of intermediate 4 (5.0 g, 22.12 mmol) in DMF (30 mL) was added (3-chloropropyl)diethylamine hydrochloride (8.23 g, 55.30 mmol) and K2CO3 (9.17 g, 66.36 mmol) at 80 °C for 3 h. The reaction was cooled, poured into an ice-water bath, and extracted with EA (2 × 200 mL). The combined organic phases were washed with brine (2 × 50 mL) and dried over Na2SO4. Upon removal of the solvent, the crude product containing the N-alkylated regioisomer was purified by FC (PE / EA = 1:10) to give intermediate 11 (1.70 g, 22.65%) as a white solid. 1 H NMR (500 MHz, CDCl3) δ 8.34 (d, J = 2.0 Hz, 1H), 8.15 (d, J = 8.0 Hz, 1H), 8.00 (dd, J = 8.0, 2.0 Hz, 2H), 4.77 (t, J = 7.0 Hz, 2H), 2.53-2.49 (m, 6H), 2.26-2.20 (m, 2H), 0.99 (t, J = 7.5 Hz, 6H). MS (ESI + ) m / z 339.1, 341.1 ([M + 1] + ).

[0250] [ka]

[0251] A mixture of Intermediate 11 (0.68 g, 2.00 mmol), Intermediate 2 (1.07 g, 3.99 mmol), potassium phosphate tribasic (0.85 g, 3.985 mmol), and Pd(dppf)Cl (0.15 g, 0.20 mmol) was suspended in 1,4-dioxane:water (12 mL, 6:1). The reaction was stirred under reflux for 16 h. The mixture was partitioned between EtOAc (2×100 mL) and water, washed with brine, dried over NaSO, and filtered. Upon removal of the solvent, the residue containing the regioisomers was purified using FC eluting with (DCM / MeOH=20 / 1) to give AKG-26 (0.54 g, 56.04%) as a gray solid. 1 H NMR (500 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.26-8.19 (m, 2H), 7.78-7.70 (m, 2H), 7.53 (dd, J= 10.5, 2.5 Hz, 1H), 5.27 (d, J = 7.5 Hz, 1H), 4.83-4.75 (m,2H), 4.17 (t, J= 11.5 Hz, 1H), 3.92 (dd, J = 11.0, 7.5 Hz, 1H), 3.74-3.69 (m, 1H), 3.62-3.58 (m, 1H), 3.51-3.28 (m, 8H), 2.14 (t, J = 8.0 Hz, 2H), 0.93 (t, J = 8.5 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 164.24, 161.03, 158.59, 154.52, 149.91, (d, J= 3.2 Hz),145.59, 141.03 (d, J = 11.8 Hz), 137.64 (d, J = 3.2Hz), 132.12, 131.40 (d, J = 4.5 Hz), 122.57, 119.06(d, J = 12.8 Hz), 114.47 (d, J = 2.8 Hz,), 105.97, 105.70, 73.96, 62.07, 51.70, 49.19, 46.75, 46.48. MS (ESI + ) m / z 470.1 ([M + 1] + ).

[0252] (18. Synthesis of AKG-27)

[0253] [ka] To a solution of intermediate 4 (6.3 g, 27.87 mmol) in DMF (42 mL) was added BocNH(CH)Br (16.6 g, 69.71 mmol) and KCO (11.1 g, 80.02 mmol) at 80 °C for 3 h. The reaction was cooled, poured into an ice-water bath, and extracted with EtOAc (2 × 200 mL). The organic phase was washed with brine (2 × 50 mL), dried over NaSO, filtered, and the solvent was evaporated under reduced pressure. The crude product, containing the N-alkylated regioisomer, was purified by FC (PE / EA = 2:1) to give intermediate 12 (14 g, 13.1%) as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.89 (d, J = 2.4 Hz, 1H), 8.28 (dd, J = 8.4 Hz, 1H), 8.11 (d, J = 8.4 Hz, 1H), 6.97 (s, 1H), 4.77 (t, J = 6.8 Hz, MS (ESI+) m / z 383.0 ([M + 1] + ).

[0254] [ka]

[0255] A solution of intermediate 12 (0.83 g, 2.15 mmol), NaHCO (0.36 g, 4.31 mmol), and intermediate 2 (1.24 g, 3.68 mmol) was suspended in 1,4-dioxane (32 mL) and water (8 mL). N was bubbled through the mixture for 5 minutes, and then Pd(dppf)Cl (0.078 g, 0.095 mmol) was added. The mixture was stirred at 90 °C for 15 hours and then cooled to RT. The mixture was partitioned between EtOAc (2 × 100 mL) and water. The organic layer was dried over NaSO, filtered, and concentrated. The filtrate was concentrated and purified by silica gel column chromatography (DCM / MeOH = 20 / 1) on silica gel to give AKG-27-1 (0.75 g; 66.9%) as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.25 (d, J = 8.5 Hz, 1H), 8.21 (d, J = 8.4 Hz, 1H), 7.78-7.70 (m, 2H), 7.54 (d, J = 8.5 Hz, 1H), 6.93 (s, 1H), 5.26 (t, J = 5.0 Hz, 1H), 4.80-4.75 (m, 3H), 4.17 (t, J = 9.0 Hz, 1H), 3.91 (t, J = 8.5 Hz, 1H), 3.71-3.69 (m, 1H), 3.60-3.59 (m, 1H), 3.06-3.03 (m, 2H), 2.15-2.12 (m, 2H), 1.37 (s, 9H) ppm. MS (ESI+) m / z 514.0 ([M + 1] + ).

[0256] [ka]

[0257] To a solution of AKG-27-1 (0.9 g, 1.75 mmol) in dry DCM (16 mL) was added HCl in dioxane (4.0 mL) at RT under a N atmosphere. The reaction mixture was stirred at the same temperature for 6 h and cooled to RT. The solvent of the reaction mixture was evaporated under reduced pressure to give AKG-27 (0.65 g, 82.5%) as a pale yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.26-8.20 (m, 5H), 7.77-7.70 (m, 2H), 7.53 (d, J = 7.6 Hz, 1H), 4.94 (d, J = 6.4 Hz, 2H), 4.77 (s, 1H), 4.51 (s, 2H), 4.16 (t, J = 8.8 Hz, 1H), 3.93 (t, J = 7.0 Hz, 1H), 3.71 (d, J = 12.4 Hz, 1H), 3.60 (d, J = 12.4 Hz, 1H), 2.94 (s, 2H), 2.34 (t, J = 6.8 Hz, 2H) ppm. MS (ESI+) m / z 414.0 ([M + 1] + ).

[0258] (19. Synthesis of AKG-28 to 31) To a solution of (R)-3-(4-bromo-3-fluorophenyl)-5-(hydroxymethyl)oxazolidin-2-one (9 g, 31 mmol) in DCM (100 mL) was added (3.92 g, 34 mmol) and TEA (3.76 g, 37 mmol). The mixture was stirred at RT for 2 h. The mixture was washed with water (2×30 mL) and brine (2×30 mL), dried over Na2SO4, filtered, and concentrated to give intermediate 13 (11.4 g, 99% yield). MS (ESI+) m / z 368 ([M + 1] + ).

[0259] To a solution of intermediate 13 (11.4 g, 31 mmol) in DMF (200 mL) was added potassium 1,3-dioxoisoindolin-2-ide (6.02 g, 32 mmol). The mixture was stirred at 90° C. overnight. The mixture was cooled, poured into water (1000 mL), and stirred for 0.5 h. The precipitate was collected and dried under reduced pressure to give intermediate 14 (11 g, 85% yield). MS (ESI+) m / z 419 ([M + 1] + ).

[0260] To a solution of intermediate 14 (11 g, 26.3 mmol) in EtOH (150 mL) was added NHNH-H0 (85%, 7.7 g, 131 mmol). The mixture was stirred at 90 °C overnight. The mixture was filtered and rinsed with EtOH (2 x 50 mL). The filtrate was concentrated to give intermediate 15 (7.6 g, 100% yield). MS (ESI+) m / z 289 ([M + 1] + ).

[0261] To a solution of intermediate 15 (7.6 g, 26.4 mmol) in THF (50 mL) and water (50 mL) was added (Boc)O (6.9 g, 32 mmol) and KCO (7.29 g, 52.8 mmol), and the mixture was stirred for 2 h. The mixture was diluted with water (100 mL) and extracted with EtOAc (3×50 mL). The combined organic extracts were washed with brine (2×50 mL), dried over NaSO, filtered, and concentrated. The residue was purified by FC (Biotage, 80 g silica gel column @ 65 mL / min, eluting with 0–60% EtOAc in petroleum ether for 30 min) to give intermediate 16 (7.8 g, 75% yield). MS (ESI+) m / z 411 ([M + 23] + ).

[0262] A mixture of intermediate 16 (7.8 g, 20 mmol), bis(pinacolato)diboron (6.54 g, 30 mmol), and KOAc (2.94 g, 30 mmol) in dioxane (100 mL) was purged with Ar for 10 min, followed by the addition of (PhP)PdCl (1.06 g, 1.5 mmol). The mixture was purged with Ar again and stirred at 90 °C overnight. The mixture was cooled, diluted with water (300 mL), and extracted with EtOAc (3 × 100 mL). The combined extracts were washed with brine (2 × 50 mL), dried over NaSO, filtered, and concentrated. The residue was purified by FC (Biotage, 80 g silica gel column @ 65 mL / min, eluting with 0–60% EtOAc in petroleum ether for 30 min) to give intermediate 18 (6.2 g, 70% yield). MS (ESI+) m / z 459 ([M + 23] + ).

[0263] Procedure C: A mixture of one of Intermediates 5 / 8 / 9 / 10 / 11 (1.0 equiv.), one of Intermediates 18 / 19 (1.5 equiv.), Pd(dppf)Cl2DCM (0.1 equiv.), and K3PO4 (2.0 equiv.) in dioxane / HO (10:1, 0.06 M) was purged with N2 and stirred at 90 °C overnight. The mixture was diluted with EtOAc, washed with water and brine, dried over anhydrous magnesium sulfate, filtered, and concentrated. The residue was purified by FC to give one of compounds AKG-28-1 / AKG-29-1 / AKG-30-1 / AKG-31-1 / AKG-38 / AKG-39 / AKG-40.

[0264] To a solution of one of compounds AKG-28-1, AKG-29-1, AKG-30-1, or AKG-31-1 in DCM (1 mL / 100 mg) was added 3N HCl (20 equiv.) in EtOAc. The mixture was stirred for 2 hours and then filtered. The solid was dried under reduced pressure or lyophilized to give one of the final compounds AKG-28, AKG-29, AKG-30, or AKG-31 (35-44% yield for the two steps).

[0265] [ka]

[0266] Use procedure C. The product was obtained from Intermediate 5 and Intermediate 18 as a white solid (0.35 g, 35% yield). 1 H NMR (500 MHz, DMSO-d6) δ 10.53 (s, 1H), 8.97 (s, 1H), 8.37 (s, 3H), 8.29 (d, J = 8.5 Hz, 1H), 8.24 (d, J = 8.5 Hz, 1H), 7.80 (t, J = 8.5 Hz, 1H), 7.69 (dd, J = 13.5, 2.0 Hz, 1H), 7.50 (dd, J = 8.5, 2.0 Hz, 1H), 5.31 (t, J = 6.0 Hz, 2H), 5.04-4.99 (m, 1H), 4.28 (t, J = 9.0 Hz, 1H), 3.96 (dd, J = 9.0, 6.5 Hz, 1H), 3.84 (s, 2H), 3.28 (s, 2H), 2.87 (s, 6H) ppm. 13 C NMR (126 MHz, D2O) δ 163.90 (s), 160.30 (s), 158.33 (s), 154.86 (s), 148.55 (s), 142.64 (s), 138.93 (d, J = 11.0 Hz), 137.74 (s), 132.43 (s), 130.39 (s), 122.46 (s), 119.03 (s), 114.36 (s), 106.41 (s), 106.18 (s), 70.31 (s), 55.23 (s), 48.07 (s), 47.69 (s), 43.29 (s), 42.19 (s) ppm. MS (ESI+) m / z 427.1 ([M + 1] + ).

[0267] [ka]

[0268] Easy to use. The resulting product, Intermediate 8, and Intermediate 18, were obtained as a white solid (0.4 g, 44% yield). 1 H NMR (400 MHz, DMSO-d6) δ 8.97 (s, 1H), 8.57-8.41 m, 6H), 8.29-8.13 (m, 2H), 7.80 (t, J = 9.0 Hz, 1H), 7.69 (dd, J = 13.5, 2.5 Hz, 1H), 7.49 (dd, J = 8.5, 2.0 Hz, 1H), 5.12 - 5.03 (m, 3H), 4.28 (t, J = 9.0 Hz, 1H), 4.02-3.98 (m, 1H), 3.54-3.51 (m, 2H), 3.33-3.26 (m, 2H). 8.28 (s, 1H), 7.73 - 7.65 (m, 1H), 7.60 (dd, J = 13.6, 2.1 Hz, 1H), 7.54 (t, J = 8.9 Hz, 1H), 7.41 (dd, J = 8.6, 2.1 Hz, 1H), 6.89 (d, J = 9.0 Hz, 1H), 5.25 (t, J = 5.6 Hz, 1H), 4.78 - 4.68 (m, 1H), 4.33 (d, J = 13.0 Hz, 2H), 4.12 (t, J = 9.0 Hz, 1H), 3.87 (dd, J = 8.9, 6.2 Hz, 1H), 3.74 - 3.64 (m, 1H), 3.62 - 3.52 (m, 1H), 2.87 - 2.71 (m, 2H), 2.23 (t, J = 7.3 Hz, 2H), 2.11 (s, 6H), 1.72 (d, J = 11.5 Hz, 2H), 1.64 - 1.49 (m, 1H), 1.34 (dd, J = 14.3, 7.0 Hz, 2H), 1.18 - 1.04 (m, 2H) ppm. 13C NMR (101 MHz, D2O) δ161.52, 160.59, 158.12, 154.86, 145.70, 141.05, 140.16, 139.65, 133.57, 130.44, 123.64, 117.70, 114.54, 106.50, 106.22, 70.34, 50.81, 47.68 ppm. MS (ESI+) m / z 399.2 ([M + 1] + ).

[0269] [ka]

[0270] Use procedure C. The product was obtained from Intermediate 10 and Intermediate 18 as a white solid (0.36 g, 40% yield). 1 H NMR (400 MHz, DMSO-d6) δ 10.94 (s, 1H), 8.96 (s, 1H), 8.52 (s, 3H), 8.28-8.22 (m, 2H), 7.79 (t, J = 8.8 Hz, 1H), 7.69 (dd, J= 13.6, 2.0 Hz, 1H), 7.49 (dd, J = 8.8, 2.0 Hz, 1H), 5.08-5.01 (m, 1H), 4.93 (t, J = 6.8 Hz, 2H), 4.28 (t, J= 9.2 Hz, 1H), 4.00 (dd, J = 9.2, 6.8 Hz, 1H), 3.29 - 3.26 (m, 2H), 3.21-3.16 (m, 2H), 2.75 (d, J = 4.8 Hz, 6H), 2.49-2.43 (m, 2H) ppm. 13C NMR (101 MHz, D2O) δ 162.39 (s), 160.58 (s), 158.11 (s), 154.88 (s), 147.20 (s), 141.66 (s), 139.28 (d, J = 11.3 Hz), 132.86 (s), 130.45 (s), 122.90 (s), 118.44 (d, J = 12.0 Hz), 114.44 (s), 106.46 (s), 106.17 (s), 70.30 (s), 54.56 (s), 50.62 (s), 47.68 (s), 42.89 (s), 42.16 (s), 23.74 (s) ppm. MS (ESI+) m / z 441 ([M + 1] + ).

[0271] [ka]

[0272] Use procedure C. The product was obtained from Intermediate 11 and Intermediate 18 as a white solid (0.36 g, 42% yield). 1 H NMR (400 MHz, DMSO-d6) δ 10.10 (s, 1H), 8.96 (s, 1H), 8.390- 8.21 (m, 5H), 7.80 (t, J= 8.8 Hz, 1H), 7.69 (dd, J = 13.6, 2.0 Hz, 1H), 7.50 (dd, J = 8.8, 2.0 Hz, 1H), 5.04-4.97 (m, 1H), 4.94 (t, J= 6.8 Hz, 2H), 4.28 (t, J = 9.2 Hz, 1H), 3.94 (dd, J = 9.6, 6.4 Hz,, 1H), 3.31-3.26 (m, 2H), 3.21-3.17 (m, 2H), 3.15-3.12 (m, 4H), 2.46-2.42 (m, 2H), 1.21 (t, J = 7.2 Hz, 6H) ppm. 13C NMR (101 MHz, D2O) δ 163.04 (s), 160.53 (s), 158.07 (s), 154.84 (s), 147.95 (d, J = 5.4 Hz), 142.36 (s), 139.05 (d, J = 11.3 Hz), 138.32 (s), 132.44 (s), 130.38 (d, J = 4.0 Hz), 122.50 (s), 118.72 (d, J = 12.8 Hz), 114.35 (s), 106.37 (s), 106.09 (s), 70.29 (s), 50.65 (s), 48.55 (s), 47.64 (d, J = 7.4 MS (ESI+) m / z 469 ([M + 1] + ).

[0273] To a solution of intermediate 15 (7.6 g, 26.4 mmol) in DCM (150 mL) was added triethylamine (TEA, 4.57 g, 6.27 mL, 52.77 mmol, 2.0 equiv), followed by acetyl chloride (AcCl, 2.6 g, 2.74 mL, 39.58 mmol, 1.5 equiv) and 4-N,N-dimethylaminopyridine (DMAP, 0.028 g, 2.64 mmol, 0.01 equiv) under N at 0-5 °C. The resulting reaction mixture was then stirred at 0-5 °C for 2 h. When TLC and LCMS showed the reaction was complete, the reaction mixture was quenched with HO (100 mL). After separation of the two layers, the aqueous layer was extracted with CHCl (2×50 mL), and the combined organic extracts were washed with HO (2×100 mL) and saturated aqueous NaCl (100 mL), dried over MgSO, and concentrated under reduced pressure. The residue was purified by FC (Biotage, 80 g silica gel column @ 65 mL / min, eluting with 0-60% EtOAc in petroleum ether for 30 min) to give intermediate 17 (6.5 g, 75% yield). MS (ESI+) m / z 332 ([M + 1] + ).

[0274] To a solution of intermediate 17 (6.5 g, 19.7 mmol) in 1,4-dioxane (100 mL) was added 1,1'-bis(diphenylphosphino)ferrocene-palladium(II) dichloride dichloromethane complex (1.61 g, 1.97 mmol), bis(pinacolato)diboron (10 g, 39.39 mmol), and KOAc (4.83 g, 49.24 mmol). The resulting reaction was stirred at 90 °C for 4 h. When TLC and LCMS indicated the reaction was complete, the reaction mixture was cooled to RT and then treated with water (100 mL) and EtOAc (100 mL). The two layers were separated, and the aqueous layer was extracted with EtOAc (2 × 50 mL). The combined organic extracts were washed with water (2 × 50 mL) and saturated aqueous NaCl (50 mL), dried over MgSO4, and concentrated under reduced pressure. The residual brown oil was purified by FC (Biotage, 80 g silica gel column @ 60 mL / min, eluting with 0-100% EtOAc in petroleum ether for 30 min) to give intermediate 19 (6.6 g, 88.7% yield). MS (ESI+) m / z 379 ([M + 1] + ).

[0275] (20. Synthesis of AKG-38~40)

[0276] [ka] Use procedure C. The product was obtained from Intermediate 5 and Intermediate 19 as a white solid (0.48 g, 60% yield). 1H NMR (400 MHz, DMSO) δ 8.95 (s, 1H), 8.29-8.19 (m, 3H), 7.77 (t, J = 8.8 Hz, 1H), 7.69 (dd, J= 13.6, 2.0 Hz, 1H), 7.50 (dd, J = 8.8, 2.0 Hz, 1H), 4.89 (t, J = 6.0 Hz, 2H), 4.81-4.76 (m, 1H), 4.20 (t, J= 9.2 Hz, 1H), 3.82 (dd, J = 9.2, 6.8 Hz, 1H), 3.45 (t, J = 5.6Hz, 2H), 2.90 (t, J = 6.0 Hz, 2H), 2.19 (s, 6H), 1.85 (s, 3H) ppm. 13 C NMR (101 MHz, DMSO-d6) δ 170.51 ,164.17 , 154.46 , 149.94 , 145.62 , 140.90, 137.63, 132.07 , 131.39 , 122.59 , 119.25 , MS (ESI+) m / z 469.2 ([M + 1] + ).

[0277] [ka]

[0278] Use procedure C. The product was obtained from Intermediate 9 and Intermediate 19 as a white solid (0.35 g, 40% yield). 1H NMR (400 MHz, DMSO) δ 8.95 (s, 1H), 8.29-8.19 (m, 3H), 7.76 (t, J = 8.8 Hz, 1H), 7.69 (dd, J = 13.6, 2.0 Hz, 1H), 7.50 (dd, J = 8.8, 2.0 Hz, 1H), 4.84-4.76 (m, 3H), 4.21 (t, J = 9.2 Hz, 1H), 3.82 (dd, J = 9.2, 6.4 Hz, 1H), 3.46 (t, J = 5.6 Hz, 2H), 3.04 (t, J = 5.6 Hz, 2H), 2.50-2.47 (m, 4H), 1.85 (s, 3H), 0.87 (t, J = 7.2 Hz, 6H) ppm. 13 C NMR (101 MHz, DMSO-d6) δ 170.50, 164.11, 154.46, 149.91, 145.68, 137.68, 132.04, 131.40, 122.53, 119.27 (d, J= 13.3 Hz), 114.65, 106.18, 105.90, 72.34, 52.11, 51.61, 47.67, 46.83, 41.87, 40.63, 40.42, 40.22, 40.01, 39.80, 39.59, 39.38, 22.92, 12.28 ppm.MS (ESI+) m / z 497 ([M + 1] + ).

[0279] [ka]

[0280] Use procedure C. The product was obtained from Intermediate 11 and Intermediate 19 as a white solid (0.36 g, 50% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.95 (s, 1H), 8.31 - 8.18 (m, 3H), 7.77 (t, J = 8.8 Hz, 1H), 7.69 (dd, J = 13.6, 2.0 Hz, 1H), 7.50 (dd, J = 8.8, 2.0 Hz, 1H), 4.88 - 4.71 (m, 3H), 4.20 (t, J = 9.2 Hz, 1H), 3.81 (dd, J = 9.2, 6.4 Hz, 1H), 3.45 (t, J = 5.6 Hz, 2H), 2.45 (s, 6H), 2.14 (s, 2H), 1.85 (s, 3H), 0.93 (s, 6H) ppm. 13 C NMR (101 MHz, DMSO-d6) δ 137.70, 131.45, 114.69, 46.79, 41.86, 40.64, 40.43, 40.22, 40.01, 39.80, 39.59, 39.38 ppm. MS (ESI+) m / z 511 ([M + 1] + ).

[0281] [Example 2] Assay for in vitro activity against Mycobacterium tuberculosis The broth microdilution MIC method used is described in Collins et al., 1997 and Gruppo et al., 2006. MIC or minimum inhibitory concentration of a compound that prevents visible growth of bacteria after overnight incubation.

[0282] Briefly, MICs were determined by a broth microdilution assay using the Alamar blue endpoint (MABA) as described by Collins et al., 1997 (Collins L, Franzblau SG (1997). Microplate alamar blue assay versus BACTEC 460 system for high-throughput screening of compounds against Mycobacterium tuberculosis and Mycobacterium avium. AAC. 41(5):1004-1009) and Gruppo et al., 2006 (Gruppo V, Johnson CM, Marietta KS, Scherman H, Zink EE, Crick DC, Adams LB, Orme IM, Lenaerts AJ. (2006) Rapid microbiologic and pharmacologic evaluation of experimental compounds against Mycobacterium tuberculosis. AAC 50:1245-1250). MABA is a 96-well colorimetric assay in which the redox indicator Alamar blue changes color from blue to pink in the presence of mycobacterial growth activity in broth medium.

[0283] Briefly, 7H9 complete medium was prepared by adding 4.7 g of Middlebrook's 7H9 broth powder (Millipore Sigma catalog #M0178), 2 mL of glycerol, and 898 mL of purified water to a 1 L flask with mixing until dissolved, followed by the addition of 100 mL of ADC solution (6 g of bovine serum albumin, 2 g of dextrose, and 3 mg of catalase dissolved in 100 mL of water) to the same 1 L flask. Compounds were brought to a concentration of 10 mg / mL in DMSO and then further diluted with DMSO to 80 μg / mL, or 40-fold the desired starting concentration of 2 μg / mL. Nine 1:2 dilution series were prepared by adding 50 μl of drug solution in the first well to 50 μl of DMSO in the next well and carrying this process forward for the next eight wells in the drug preparation plate. Stocks of M. tuberculosis (M.tb) H34Rv and M.tb Erdman strains were cultured at 3–4 x 10 7 From their initial concentration of CFU / mL, 5x10 5 Dilutions were made to a final concentration of CFU / mL and mixed thoroughly by pipetting up and down using a multichannel pipettor.

[0284] 5x10 5Assay plates were prepared by transferring 100 μl of medium inoculated with CFU / mL to all wells. Subsequently, 2.5 μL of each drug dilution from the drug preparation plate was transferred to the corresponding well in the assay plate. The assay plates were then placed in Ziploc® bags and placed inside an incubator, where they were incubated at 37°C. The plates were then read at OD600nm on a plate reader on days 3 and 10. After the OD600 reading on day 10, 10 μl of Alamar Blue dye was added to each analytical well. On day 12, all assay plates were scanned on a flatbed color scanner. The lowest serial antimicrobial concentration (typically 2-fold serial dilutions) that did not produce a visible color change from blue to pink with Alamar Blue and / or showed an 80% or greater reduction in OD600 relative to drug-free control wells was considered the MIC for these compounds.

[0285] Assays were performed using two unique drug-susceptible strains (M.tb Erdman and M.tb H37Rv). MIC assays can also be performed in the presence of 4% (w / v) human serum albumin (huSA) (Sigma #A1653) to assess potential protein binding (serum shift assay). Generally, a shift in the MIC between two wells (a four-fold shift in the MIC) is considered significant. For PA-824 (positive control), a four-fold shift in the MIC is expected.

[0286] MICs were measured by the Alamar Blue (MABA) readout or by the optical density readout (OD600), which matched or differed by only one two-fold dilution, within the limits of the assay. All compounds tested showed consistent MIC values ​​against both Mtb Erdman and H37Rv, or were within one two-fold dilution, except for one compound, AKG-40, which showed higher MIC values ​​of 1-2 μg / mL against Erdman and an MIC of 0.5 against H37Rv. This difference may be due to slower growth (lower OD reading) on ​​Erdman plates.

[0287] Linezolid showed predicted MIC values ​​of 2 μg / mL, tedizolid showed 0.25 μg / mL, and bedaquiline showed 0.125 μg / mL. These values ​​are consistent with previous MIC data and published values ​​(Ruiz et al. Antimicrob. Agents Chemother. 2019, Mar 27;63(4),pii:e01939-18; Reddy et al. Antimicrob Agents Chemother. 2010 Jul;54(7):2840-6; Torrea et al. J Antimicrob Chemother. 2015 Aug;70(8):2300-5). AKG-28 showed significantly greater activity than tedizolid, with MICs ranging from 0.03 to 0.015 μg / mL. Among oxazolidinone analogs containing an acetamide group, AKG-39 showed an MIC of 0.5 μg / mL, AKG-40 showed MICs ranging from 1 to 0.5 μg / mL, and AKG-38, with an MIC of 0.06 μg / mL, also showed several-fold higher activity than tedizolid.

[0288] Molecules bearing an amine or acetamide group at the C5 position of the oxazolidinone were more active (AKG-3 vs. tedizolid, AKG-28 or AKG-38 vs. AKG-16, AKG-39 vs. AKG-24, AKG-40 vs. AKG-26), while compounds bearing an aminoalkyl side chain on the tetrazole showed favorable activity. Substitution of the t-butoxycarbonylamino (Boc-NH) group at the C5 position of the oxazolidinone with a primary amine (AKG-28-1 vs. AKG-28) or acetamide (AKG-28-1 vs. AKG-38) resulted in decreased activity. Compounds containing dimethylaminoalkyl side chains were particularly superior when compared with aminoethyl or diethylaminoethyl analogs (AKG-16 vs. AKG-24, AKG-28 vs. AKG-29, AKG-30 vs. AKG-31). Similarly, shorter dialkylaminoalkyl side chains on the tetrazole ring (e.g., ethylene vs. propylene) showed greater activity (AKG-16 vs. AKG-25, AKG-24 vs. AKG-26, AKG-28 vs. AKG-30). Analogs with substitutions on the 2'-position of the tetrazole were more active than those with substitutions at the 1'-position (AKG-16 vs. AKG-21, AKG-23 vs. AKG-22).

[0289] [ka]

[0290] [Table 8]

[0291] [Example 3] [In vitro cytotoxicity assays against human kidney and human hepatocytes] Compounds were tested in vitro across a 10-fold dilution series to determine IC50 in African green monkey kidney (Vero; ATCC #CCL81) or human hepatocyte / liver (HepG2; ATCC #HB8065) ​​cells. Because these molecules are generally expected to be non-toxic, a doxorubicin positive control was included in all studies. Data are reported as whole cell survival curves, as well as calculations of actual IC50 values ​​for each compound.

[0292] Adherent cells were grown to approximately 80% confluence. After trypsinization by adding 0.25% trypsin-EDTA (Gibco #25200-072), the cells were spun down, and 5 ml of growth medium (MEM medium; Corning #10010CM) was added to disperse the cells. Cell density was determined using a hemocytometer. Growth medium (MEM medium containing 10% FBS; Corning #35015CV) was added to the cells to adjust the appropriate cell concentration. Then, 200 μl of cells (5,000 cells / well) were added to a 96-well clear flat-bottom plate (Costar #9804) and incubated in the plate at 37°C in a humidified incubator with 5% CO2 for 24 hours.

[0293] Serial dilutions of test compounds were prepared using growth medium as the solvent (Table 2). These compounds were provided as sterile aqueous solutions of the HCl salt with a concentration of 5 mg / ml. To make the dilutions, each drug stock was warmed to room temperature, vortexed, and visually inspected for sediment. If solid drug was present, the stock was heated in a 60°C water bath and then allowed to cool to near room temperature. Based on the treatment concentration, 20-fold diluted standard stocks were made by serial dilution. These were further diluted 1-fold in growth medium, and the highest drug concentration tested was 250 μg / ml.

[0294] Compounds were added to wells in a 1:2 dilution series starting from an initial concentration of 250 μg / ml for each compound by aspirating the old medium and replacing it with 200 μl of drug-containing medium. Plates were incubated for 72 hours at 37°C in a humidified incubator with 5% CO2. At the end of the compound incubation period, the medium in each well was replaced with 100 μl of 1X PrestoBlue Cell Viability Reagent (ThermoFisher catalog #A13261). Plates were incubated for 30 minutes to 2 hours at 37°C in a humidified incubator with 5% CO2. Readings were taken at 30, 60, and 120 minutes. Fluorescence was read at 560 nm excitation and 590 nm emission using a SpectraMax M5 plate reader (Molecular Devices). Background was corrected by subtracting the RFU of a control containing culture medium only (background control well) from all sample readings. Calculate the percentage of cytotoxicity using the following formula: Cytotoxicity %=[(RFU .Medium -RFU Treatment ) / RFU .Medium ]×100%.

[0295] The IC50 was determined using GraphPad Prism using the following formula: Y=100 / (1+10^((LogIC50-X) * Hill gradient))).

[0296] [Table 9]

[0297] Surprisingly, many of the analogs containing a hydroxyl group on the C5 side chain of the oxazolidinone ring, which mimics the substituent of the active metabolite tedizolid for tedizolid phosphate, exhibited the highest hepatotoxicity, exhibiting single-digit IC50s in the HepG2 hepatocyte cell line. Tedizolid is the most active oxazolidinone currently approved for the treatment of MRSA and shares structural similarity with the compounds described herein in the tetrazole D ring, pyridyl C ring, and aryl B ring (see Figure 6). However, increased toxicity to hepatocytes results in a relatively low selectivity index for the oxazolidinones with a hydroxyl on the C5 side chain (AKG-23, AKG-25, AKG-26, and AKG-27) compared with those with an amino or acetamido group at the same position on the C5 side chain (AKG28-31, AKG38-40, and AKG-3).

[0298] [Example 4] [Determination of selectivity index] Selectivity indices (SI) were calculated to determine the relative inhibitory activity of compounds against two M. tuberculosis strains, Erdman and H37Rv, compared to that against mammalian cells, i.e., African green monkey kidney (VERO) or human hepatocyte-derived (HepG2) cells, as described in Examples 2 and 3, respectively. A high SI is preferred because it indicates favorable killing of the M. tuberculosis strain at a drug concentration that is less harmful to normal cells in the body. The selectivity index was calculated using the following formula: SI=IC 50,mammalian / MIC bacteria (wherein the bacteria is Mycobacterium tuberculosis, either the Erdman or H37Rv strain, and the mammalian cells are the VERO or HepG2 cell lines).

[0299] If the IC50 was higher than the highest value tested for VERO or HepG2 cells, the SI was indicated as higher (>) than the ratio calculated using that highest concentration. Similarly, if the MIC for the Erdman or H37Rv strains was higher than the highest concentration of drug tested (8 μg / ml), the SI was indicated as lower (<) than the ratio calculated using that highest concentration. If both numbers were above the highest concentration tested, the calculated value was indicated as not determined (nd). The results are shown in Table 4. The SI did not directly correlate with the activity of the molecule in either the M. tuberculosis strains or the mammalian cell lines, and increased potency in M. tuberculosis strains did not directly correlate with increased toxicity to the mammalian cell lines. For example, AKG-38 exhibited nanomolar MICs against both M. tuberculosis strains, but it was relatively inactive against both VERO and HepG2 cell lines compared to the other molecules in the panel, giving a high SI. This was also observed for AKG-28. It is noteworthy that both the AKG-28 and AKG-38 molecules had a dimethylaminoethyl substituent at the 2' position of the tetrazole ring.

[0300] [Table 10]

[0301] In some embodiments, the compound of interest has a saturation level of greater than 100, greater than 200, greater than 300, greater than 400, greater than 500, greater than 1000, greater than 1500, greater than 2000, greater than 2500, greater than 3000, greater than 3500, greater than 4000, greater than 4500, greater than 5000, greater than 5500, greater than 6000, greater than 6500, 100-7000, 100-6000, 100-5000, 100-4000, 100-3000, 100-2000, 100-1000, 100-900, 100-800, 100-700, 100-600, 100-500, 100-400, 100-400, 100-500, 100-400, 100-500, 100-400, 100-6500, 100-7000, 100-800, 100-700, 100-600, 100-500, 100-400, 100-500, 100-400, 100-6500, 100-7000, 100-700, 100-600, 100-500, 100-400, 100-6500, 100-7000, 100-800, 100-800, 100-900, 100-900, 0~300, 100~200, 200~7000, 200~6000, 200~5000, 200~4000, 200~3000, 200~2000, 200~1000, 200~900, 200~800, 200~700, 200~600, 200~500, 200~400, 200~300, 300~700 The compound of interest has an SI index with respect to Erd / HepG2 and H37Rv / HepG2 of 0, 300-6000, 300-5000, 300-4000, 300-3000, 300-2000, 300-1000, 300-900, 300-800, 300-700, 300-600, 300-500, or 300-400. In some embodiments, the compound of interest has an SI index with respect to Erd / HepG2 and H37Rv / HepG2 in the range of 100-1700, 200-1700, or 300-1700.

[0302] Compounds bearing an amino or acetamido group on the C5 side chain of the oxazolidinone ring and an aminoalkyl group at the 2' position of the tetrazole ring exhibited relatively high SI compared with those bearing a hydroxyl group on the C5 side chain. Furthermore, specific tetrazole substitutions further improved SI, with dimethylaminoethyl substitution at the 2' position of the tetrazole ring being preferred over methyl, diethylaminoethyl, aminoethyl, or dimethylaminopropyl substitutions at the same position (AKG-28 and AKG-38). Moving the dimethylaminoethyl group to the 1' position of the tetrazole ring (compounds AKG-21 vs. AKG-28) unexpectedly resulted in a dramatic loss of activity against M. tuberculosis.

[0303] [Example 5] Assay for in vitro activity against methicillin-resistant Staphylococcus aureus (MRSA). The activity of lead oxazolidinone inhibitors was measured and showed sufficient potency against the Gram-positive bacterium methicillin-resistant Staphylococcus aureus (MRSA) to justify subsequent delivery in liposomal form for its treatment. In some embodiments, the MIC for two of the three strains evaluated is less than 6 μg / mL. In some embodiments, the MIC for two of the three strains evaluated is less than 2 μg / mL, with less than 2 μg / mL being more preferred.

[0304] Three S. aureus strains were grown overnight at 37°C under ambient atmosphere on trypticase soy agar plates supplemented with 5% sheep blood cells. Cultures were aseptically swabbed and transferred to tubes of sterile water, and the optical density was adjusted to 0.5 at 600 nm. Cultures were then diluted 1:100 to yield approximately 5 x 10 per well in 120 μL. 5 cells were delivered. After incubation, the MIC of the test article was determined by the presence / absence of growth in each well. MIC analysis was performed in triplicate.

[0305] Tedizolid demonstrated MICs ranging from 0.206 to 0.617 μg / ml, similar to the 0.5 μg / ml described in U.S. Patent No. 7,816,379. Interestingly, all molecules with a primary amine modification at R2 of the oxazolidinone ring (AKG-3, AKG-28, AKG-29, and AKG-30) exhibited negligible activity (>50 μg / ml) against all three MRSA strains. Molecules with an acetamide group at the same position (AKG-38, AKG-39, and AKG-40) were three to nine times less active than tedizolid itself against the three MRSA strains.

[0306] [Table 11]

[0307] [Example 6] [Liposome composition] (General Protocol) 1. The lipid components (phospholipids (PhL), cholesterol, and, if necessary, PEG-lipid derivatives and / or fluorescently labeled lipids) were mixed with a volume of 100% ethanol equal to one-tenth of the calculated volume (V) to obtain a lipid suspension containing approximately 60 mM phospholipids, and stirred at a temperature of 65-68 °C until complete dissolution of the lipids.

[0308] Neutral phospholipids include diacylphosphatidylcholine, dialkylphosphatidylcholine, sphingomyelin, and diacylphosphatidylethanolamine. Hydrogenated soybean phosphatidylcholine, distearoylphosphatidylcholine, and egg sphingomyelin are some of the preferred phospholipids.

[0309] The PEG-lipid component may comprise PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (PEG-DSPE), or N-palmitoyl-sphingosine-1-{succinyl[methoxy(polyethylene glycol)2000]} (PEG-ceramide). The molecular weight of the PEG-lipid component may also vary from 1,500 to 6,000 g / mol, but is preferably about 2,000 MW.

[0310] Lipid fluorescent labels include 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine-5,5'-disulfonic acid (DiIC18(3)-DS), 1,1'-dioctadecyl-3,3,3',3'-tetramethylindodicarbocyanine-5,5'-disulfonic acid (DiIC8(5)-DS), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-(cyanine7) (18:0 Cy7 PE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000]-N-(cyanine7) (DSPE PEG(2000)-N-Cy7), and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-(cyanine5) (18:0 Cy5 PE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000]-N-(cyanine 5) (DSPE PEG(2000)-N-Cy5), 1-oleoyl-2-[12-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]dodecanoyl]-sn-glycero-3-phosphocholine (18:1-12:0 NBD PC).

[0311] 2. The ethanolic lipid solution was mixed with volume V of scavenger solution (0.25-0.5 M ammonium sulfate or 1 N triethylammonium sucrose octasulfate) with stirring at 65-68 °C until a homogenous suspension was obtained.

[0312] Potential scavengers include, but are not limited to, diethylammonium or triethylammonium salts of sucrose octasulfate, ammonium sulfate, ammonium citrate, citric acid, dextran sulfate, polyvinyl sulfonic acid, or the ammonium salt of inositol hexaphosphate, at a concentration of 0.1 to 2 g equivalents / L (0.1 to 2 N), preferably 0.2 to 1.5 N. Typically, an ammonium salt is used, which may include ammonium itself, monoalkyl-, dialkyl-, or trialkyl-ammonium salts.

[0313] The lipid suspension was extruded at least three times through a stack of track-etched polycarbonate membranes, typically two or four membranes with a nominal pore size of 100 nm and one membrane with a nominal pore size of 200 nm (Whatman Nuclepore, USA), using a thermobarrel extrusion apparatus (Lipex, Canada) at 400-450 psi and 65-68°C. When two 100 nm membranes were used in a 100 ml Lipex extrusion apparatus, the extrusion pressure was typically 260-300 psi. The resulting liposomes had a Z-average particle size (diameter) Xz of approximately 80 to approximately 130 nm and a PDI of less than 0.1.

[0314] 4. The extruded lipid suspension (known to contain unilamellar and / or oligolamellar liposomes) was chilled in a refrigerator (2-8°C) and filtered under positive pressure through a 0.2 μm polyethersulfone (PES) membrane filter.

[0315] 5. An aliquot of the thus-prepared, extruded, and filtered liposome suspension was chromatographed on a gravity-fed Sepharose CL-4B size-exclusion column (eluent: Type 1 water) to purify the liposomes from extraliposomal entrapment agents. The purified liposomes were collected near the void volume fraction of the column. For scale-up studies, this step was performed using tangential flow filtration (TFF) on a hollow fiber cartridge (Repligen Spectrum MicroKros PS or mPES membrane with a 500 kDa MWCO) with 8-10 volume exchanges with Type 1 or USP "Water for Injection" endotoxin-free water (or until the conductivity of the liposome suspension fell below 200 μS / cm).

[0316] 6. Lipid concentrations in purified, extruded liposome preparations were determined using HPLC with UV detection by measuring cholesterol concentrations and correcting for known phospholipid / cholesterol molar ratios. Alternatively, phospholipid content was directly quantified using a spectrophotometric phosphomolybdenum blue method.

[0317] 7. Drugs were dissolved in type 1 or endotoxin-free purified water in the form of hydrochloride salts at concentrations of 5–20 mg / ml (e.g., AKG-3 and AKG-5 were used as monohydrochlorides, and AKG-28 and AKG-29 were used as dihydrochlorides). To drugs prepared in free base form (e.g., AKG-16, AKG-38), an equal volume of HCl was added. If necessary, the pH of the solution was adjusted to 2.5–5.5 using 1N NaOH, HCl, or tris(hydroxymethyl)aminomethane (Tris) base solution, and the solution was filtered through a 0.2 μm PES filter under positive pressure. Where necessary, the drug concentration in the stock solution was verified by HPLC with UV detection at 305 nm.

[0318] 8. The purified liposomes and drug stock solution from step 5 were mixed in the presence of the amount of osmotic agent (typically dextrose) and water necessary to provide the desired drug:phospholipid (DL) ratio, a drug concentration ranging from 1.5 to 3.3 mg / ml, and an osmolality equal to the measured osmolality of the sequestering agent solution from step 2. If necessary, a buffer solution of the desired pH (typically, pH 4 to pH 7) was added. In some cases, the amount of osmotic agent added (e.g., approximately 45 g / L of dextrose) resulted in an osmolality lower than the measured osmolality of the sequestering agent solution, and loading was performed with 6 to 8 mg / ml of drug.

[0319] 9. The drug-liposome mixture was incubated at 65-68°C for approximately 15-20 minutes with constant stirring and then rapidly cooled on ice. After 5-10 minutes, the mixture was allowed to reach ambient temperature and adjusted to 0.1 M NaCl by adding the calculated amount of 3 M NaCl stock solution.

[0320] 10. Drug-loaded liposomes were purified from unencapsulated drug by size-exclusion chromatography (SEC) on a gravity-fed Sepharose CL-4B column, eluting with 10 mM HEPES buffer, pH 7.0 (HBS-7) in 140–144 mM NaCl. The liposome fraction was collected near the column void volume. For scale-up studies, purification and buffer exchange were performed using TFF as described under item 5 above, with a 10-fold volume exchange with HBS-7 buffer. In scale-up processes, approximately 8-fold volume exchanges were typically used. If necessary, purified liposomes were concentrated by continuing the TFF process without buffer feeding. The purified, drug-loaded liposomes were aseptically filtered using a 0.2 μm sterile PES filter under positive pressure and stored in a refrigerator (2–8°C).

[0321] 11. Drug and lipid concentrations in purified drug-loaded liposome preparations were determined by HPLC. Alternatively, a spectrophotometric (phosphomolybdenum blue) method was used for phospholipid quantification, and drug was quantified by UV absorption (302-305 nm) in liposome samples solubilized in 70% isopropanol-0.1 N HCl in the presence of 6.5 mg / ml sodium dodecyl sulfate. Encapsulation efficiency was assessed by: EE, %=DL / DL0 * 100% (where DL is the drug to phospholipid ratio in the liposome-loaded mixture before SEC or TFF purification, and DL is the drug to phospholipid ratio in the drug-loaded liposomes after purification (step 10).) It was decided as follows.

[0322] 12. The mean liposome size (Z-average diameter, Xz) and polydispersity index (PDI) were determined using dynamic laser scattering by the method of cumulants on a Zetasizer mu-V, Zetasizer Nano, or Zetasizer Pro (Malvern Panalytical, US).

[0323] [Example 7] [In vivo stability and blood clearance of liposomes] The drug encapsulation stability and blood clearance rate of liposomes encapsulating compounds according to the present disclosure were tested in mice according to the following general protocol. Three groups of mice (C3H female or CD-1 male) of a given laboratory strain were injected with drug-loaded liposomes via the tail vein at a dose of 9 mg of drug per kg of body weight. At time points 1 and 2, blood was collected from the retro-orbital sinus and the animals were sacrificed. Typically, blood collection time points included 5 minutes, 1 hour, 6 hours, and 24 hours after injection. Plasma was separated by centrifugation, extracted with acidified isopropanol containing a solubilizing agent (sodium octasulfate) as needed, and analyzed for drug and lipids (when liposomes contained the lipid label, DiIC18(3)-DS) by HPLC. Blood clearance of liposomal drugs was expressed as the percentage of the injected dose remaining at a given time point. The in vivo stability of drug encapsulation was assessed by the percent change (decrease) in the DL ratio in plasma at a given time point compared to the DL value before injection.

[0324] [Example 8] Loading of AKG-3, AKG-5, and AKG-16 into liposomes at different pH levels. A trimethylammonium sucrose octasulfate scavenger solution was prepared by passing a solution of commercially available potassium sucrose octasulfate (40.2 g in 145 ml of water) through a 500 ml ion exchange column of hydrogen-form Dowex 50Wx8 100-200 mesh and titrating the resulting free acid form of sucrose octasulfate with neat triethylamine to pH 6.2. The concentration of triethylammonium sucrose octasulfate (TEA-SOS) (1 N, corresponding to 0.125 M sucrose octasulfate) was estimated from the amount of triethylamine consumed in the titration. The remaining potassium was estimated using a Horiba LAQUATwin K-11 potassium analyzer by the addition method and was found to be less than 0.1% of the initial potassium amount.

[0325] Liposomes (0.5 mol% HSPC) composed of hydrogenated soybean phosphatidylcholine (HSPC) (Lipoid, Germany), cholesterol (3:2 molar ratio), and methoxypoly(ethylene glycol) ether of 1,2-distearoylglycerol (PEG-DSG, PEG molecular weight 2000, NOF, Japan) with 1N trimethylammonium sucrose octasulfate (TEA-SOS) as a sequestering agent were prepared essentially as described in the general protocol above. The drug loading step was carried out at a pH of 500 mg / mol DL (DL0) in the presence of 16 mM morpholinoethanesulfonic acid (MES)-4 mM sodium citrate buffer with a pH ranging from 4.3 to 7.1, as well as without the addition of any buffering substance (pH 5.2-5.9). All drugs were encapsulated in liposomes with high efficiency over the entire pH range tested (except for AKG-16 at pH 4.38, which was loaded with 93.3% efficiency, exceeding 98%) (Figure 1). Addition of buffering substances was not necessary for efficient encapsulation.

[0326] [Example 9] Encapsulation of AKG-3, AKG-5, and AKG-16 in liposomes with TEA-SOS scavengers at different DL ratios. Liposomes composed of HSPC, cholesterol (3:2 molar ratio), and PEG-DSG (0.5 mol% of HSPC) with 1N TEA-SOS as the sequestering agent were prepared essentially as described in the general protocol (Example 6). The drug loading step was carried out without added buffer (pH 4.98-6.22) at DL ratios ranging from 750 to 1500 g / mol pHL. Maximum drug loadings for compounds 3, 5, and 16 were observed in the ranges of 900-930 g / mol pHL, 982-1197 g / mol pHL, and 938-951 g / mol pHL, respectively, with loading efficiencies at or near maximum drug loading of at least 97.6%, 96.0%, or 85.2%, respectively (Figures 2A and 2B).

[0327] [Example 10] Encapsulation of AKG-3, AKG-5, and AKG-16 in liposomes using high PEGylation degree or 0.25 M ammonium sulfate (AS) as a trapping agent. Liposomes composed of HSPC and cholesterol with various PEG-DSG contents (3:2 molar ratio) and entrapment agents were prepared according to the general protocol and loaded with compounds AKG-3, AKG-5, and AKG-16 at DL ratios of 250 or 500 g / mol PhL as described in Example 9. All three compounds were loaded into liposomes with high efficiency, as shown in Table 6 below.

[0328] [Table 12]

[0329] Thus, compounds AKG-3, AKG-5, and AKG-16 were efficiently loaded into phospholipid-cholesterol liposomes with high levels of PEGylation and ammonium sulfate as the intraliposomal drug entrapping agent. However, loading efficiency decreased for two of the three oxazolidinones (AKG-3 and AKG-16) when loaded at a higher drug:lipid ratio of 500 g drug / mol PhL using 0.25 M ammonium sulfate as the entrapping agent.

[0330] [Example 11] Loading of compounds AKG-3, AKG-5, and AKG-16 into liposomes using 0.5 M AS as a scavenger. Liposomes composed of HSPC and cholesterol (3:2 molar ratio) with 0.5 mol% or 5% mol% PEG-DSG (relative to PhL) and 0.5 M ammonium sulfate (AS) as a sequestering agent were prepared according to the general protocol and loaded with compounds AKG-3, AKG-5, and AKG-16 as described in Example 8 at DL ratios ranging from 500 to 1500 g / mol PhL. The results are shown in Figures 3A, 3B, 3C, and 3D. All three compounds were loaded into both liposomes at DL ratios of 420 to 450 g / mol PhL with encapsulation efficiencies of 93 to 100%; the maximum drug payloads were as follows:

[0331] [Table 13]

[0332] All three tested compounds were loaded into 0.5M ammonium sulfate liposomes at more than 500 g of drug / mol PhL in the preparation containing 0.5 mol% PEG-DSG, and compounds AKG-3 and AKG-16 were loaded in the formulation containing 5 mol% PEG-DSG. These high loading levels are important in achieving sufficient doses of drug administered for disease treatment. Although loading efficiency was significantly improved compared to Example 10, where loading efficiency was lower using 0.25M ammonium sulfate, this indicates that, despite higher osmolality and the possibility of osmotic burst, higher ammonium sulfate concentrations of 0.5M are improved in terms of the amount of drug that can be loaded per mole of phospholipid, and preferably for anti-infective agents, where low toxicity and high doses can lead to improved outcomes.

[0333] [Example 12] Loading of Compounds AKG-3, AKG-5, AKG-16, and AKG-28 into Liposomes of Various Compositions Containing Fluorescent Lipid Labels. Liposomes composed of HSPC and cholesterol (60:40 molar ratio) with 0.5 mol% PEG-DSG (relative to PhL), 0.15 mol% lipid fluorescently labeled DiIC18(3)-DS (ThermoFisher, USA), and 0.5 M ammonium sulfate (AS) or 1 N TEA-SOS as a scavenger were prepared according to the general protocol and loaded with compounds AKG-3, AKG-5, and AKG-16 at pH 4.7-5.8 (no buffer substances added) as described in Example 11. The liposomes had the following characteristics:

[0334] [Table 14]

[0335] All three drugs were efficiently loaded into liposomes. Degradation of AKG-5 during liposome loading was detected as the appearance of a second peak on HPLC.

[0336] Liposomes composed of various phospholipids (HSPC, distearoylphosphatidylcholine (DSPC, Avanti Polar Lipids, USA), or egg sphingomyelin (ESM, Lipoid, Germany)) and cholesterol (60:40 molar ratio) containing various amounts of PEG-DSG or N-methoxypoly(ethylene glycol)oxycarbonyl-1,2-distearoylphosphatidylethanolamine (PEG-DSPE, PEG molecular weight 2000, Lipoid, Germany), and the lipid fluorescently labeled DiIC18(3)-DS (0.15 mol% with respect to PhL) were prepared according to the same general protocol using different trapping agents and loaded with AKG-16 in a similar manner. Where indicated, the liposome extrusion step of the general protocol was supplemented with extrusion through two stacked polycarbonate membranes with 50 nm pore sizes. The liposomes had the following characteristics:

[0337] [Table 15]

[0338] Liposomes composed of HSPC and cholesterol (3:2 molar ratio) with 9.2 mol% PEG-DSPE (relative to PhL), 0.15 mol% lipid-labeled DiIC18(3)-DS, and 0.25 M ammonium sulfate (AS) as a trapping agent were prepared according to the general protocol and Example 12 using an additional 50 nm extrusion and loaded with AKG-28 at a drug:lipid ratio (DL) of 150 g / mol PhL. Liposomes (batch ID 98) have a DL ratio of 73.8 g / mol PhL, a Z-average liposome size of 77.8 nm, and a size polydispersity index (PDI) of 0.090.

[0339] These studies demonstrate that AKG-3, AKG-5, and AKG-16 can be efficiently loaded into liposomes with a range of lipid compositions, including HSPC, DSPC, or ESM as the neutral phospholipid component, or low (0.5 mol%) or high (5 mol%) PEG-lipid contents. However, compared with 1 N TEA-SOS, the efficiency significantly decreased when 0.25 M AS was used, from approximately 500 g AKG-16 / mol PhL to 128 g AKG-16 / mol PhL. A similarly low loading efficiency (i.e., 73.8 g / mol PhL) was observed when AKG-28 was loaded with 0.25 M AS. This suggests that TEA-SOS or higher concentrations of AS may be preferred for loading higher concentrations of compounds into liposomes.

[0340] [Example 13] [Blood Persistence and In Vivo Encapsulation Stability of Liposomes of Example 12 in Mice] Tests were performed on male CD-1 mice as described in the general protocol above.

[0341] [Table 16]

[0342] These studies show that liposomes composed of varying neutral phospholipid components (HSPC, DSPC, or SM) and loaded with AKG-16 using a TEA-SOS scavenger were slowly cleared, with over 30% of the injected dose remaining in plasma at 6 hours for most formulations. Furthermore, most formulations showed good drug retention, with the exception of liposome batch ID 97, which contained AKG-16 loaded using 0.25 M AS. However, the results suggest that loading the drug using 0.25 M ammonium sulfate not only resulted in low loading efficiency, as shown in Table 9, but also in a low DL ratio (3.7%) at 6 hours due to significant leakage from liposomes in this formulation.

[0343] [Example 14] Encapsulation of compounds AKG-28 and AKG-38 in liposomes with different DL ratios and various entrapment agents. Liposomes composed of HSPC and cholesterol with 0.5 mol% PEG-DSG (relative to PhL), 0.15 mol% lipid-labeled DiIC18(3)-DS, and 0.5 M ammonium sulfate (AS) or 1 N TEA-SOS as a scavenger were prepared according to the general protocol and loaded with compounds AKG-28 and AKG-38 at pH 4.95–5.17 (no buffer added) and DL ratios ranging from 300–1050 g / mol PhL (AKG-28) or 400–1400 g / mol PhL (AKG-38), as described in Example 8. When using 0.5 M AS, the maximum drug loadings for compounds AKG-28 and AKG-38 ranged from 404 to 424 g / mol PhL and 818 to 842 g / mol PhL, respectively, with loading efficiencies above 95% at drug loadings of 302 g / mol PhL (quantitative loading) and 387 to 764 g / mol PhL (95.5 to 96.7% loading), respectively. When using 1 N TEA-SOS, the maximum drug loadings for compounds AKG-28 and AKG-38 ranged from 315 to 328 g / mol PhL and 989 g / mol PhL, respectively, with loading efficiencies of 83.5% at drug loadings of 250 g / mol PhL and 400 to 777 g / mol PhL (above 97.2% loading), respectively (Figures 4A and 4B).

[0344] While AKG-38 demonstrated near-quantitative loading of 400-800 g AKG-38 / mol PhL, the resulting drug-to-lipid ratio remained flat for AKG-28 over the range of 250-1000 g AKG-28 / mol PhL, suggesting a lower maximum drug loading for AKG-28 than for AKG-38. It should be appreciated that the higher potency previously demonstrated for AKG-28 makes liposomal formulations of AKG-28 effective for the treatment of infectious diseases such as tuberculosis.

[0345] [Example 15] Encapsulation of compounds AKG-28 and AKG-38 in liposomes using different phospholipid compositions, PEGylation degrees, and entrapment agents. Liposomes composed of phospholipid (PhL) and cholesterol (3:2 molar ratio), PEG-DSG, and DiIC18(3)-DS (0.15 mol% of PhL) with 0.5 M AS or 1 N TEA-SOS as a scavenger were prepared according to the general protocol and loaded with compounds AKG-28 and AKG-38 (in the absence of added buffer) at DL ratios selected to optimize drug loading and encapsulation efficiency (EE). The results are shown in Tables 10 and 11 below.

[0346] [Table 17]

[0347] [Table 18]

[0348] This example demonstrates that AKG-28 can be efficiently loaded into liposomes composed of HSPCs using 0.5 M AS or 1 N TEA-SOS as the entrapment agent with a maximum drug loading of 230-275 g AKG-28 / mol PhL. However, formulations containing sphingomyelin as the neutral phospholipid for this compound showed relatively low loading, with a maximum of only approximately 110 g AKG-28 / mol PhL.

[0349] Compound AKG-38 was loaded at significantly higher D / L ratios of 525-600 g / mol when the drug was added at 600 g AKG-38 / mol PhL, or greater than 735 g / mol when added at 800 g AKG-38 / mol PhL, using 0.5 M AS or 1 N TEA-SOS. The loading of compound AKG-38 was less sensitive to the presence of sphingomyelin than AKG-28.

[0350] [Example 16] Encapsulation of compounds AKG-16, AKG-28, AKG-29, and AKG-38 in liposomes using high PEGylation and 0.5 M ammonium sulfate as a trapping agent. Liposomes composed of HSPC and cholesterol (3:2 molar ratio) with 0.5 M ammonium sulfate as a scavenger, PEG-DSG (5 mol%), and DiIC18(3)-DS (0.15 mol%) were prepared according to the general protocol and loaded with compounds AKG-16, AKG-28, AKG-29, or AKG-38 (in the absence of added buffer) at DL ratios selected to optimize drug loading and encapsulation efficiency (EE). The results are shown in Table 13 below.

[0351] [Table 19]

[0352] The data show that all compounds containing a dimethylaminoethyl substituent at the 2-position of the tetrazole ring were efficiently loaded into liposomes at over 80% efficiency, whereas AKG-29, which has an aminoethyl substituent at the same position, was poorly loaded into liposomes with an efficiency of 14.5% and a final drug loading of 43.6 g AKG-29 / mol PhL. This indicates that despite the presence of titratable amines in all compounds tested, compounds with an ammonium group substituted on the tetrazole ring (e.g., an N,N-dimethylaminoethyl group) allowed for more efficient drug loading than those with a primary amine (aminoethyl group) at the same position.

[0353] [Example 17] [Blood Persistence and In Vivo Encapsulation Stability of Liposomes of Examples 15 and 16 in Mice] Tests were performed on male CD-1 mice as described in the general protocol above.

[0354] [Table 20]

[0355] The data showed that the drug in liposome batches IDs 128, 132, 142, 144, and 145, all with 0.5 M AS as a sequestering agent, lost 25–60% of the encapsulated drug almost immediately upon contact with blood, as indicated by the low DL ratio at 5 min and a further decline in the DL ratio at 6 h, particularly pronounced for liposomes loaded with AKG-38 and AKG-16. Thus, formulations with 0.5 mol% or 5 mol% PEG-DSG and 40 mol% cholesterol, 0.5 M AS (as a sequestering agent), were unable to retain drug as efficiently as formulations with 1 N TEA-SOS (liposome batches IDs 129, 130, 133–135), when the % initial DL ratio at both 5 min and 6 h was above 80%.

[0356] [Example 18] Preparation and loading of AKG-28 and AKG-38 in PEGylated liposomes with varying phospholipid to cholesterol ratios. Liposomes containing 5 mol% PEG-DSG or PEG-DSPE (relative to PhL), 0.15 mol% lipid-labeled DiIC18(3)-DS, and 0.5 M ammonium sulfate (AS) or 1 N TEA-SOS as a scavenger were prepared according to the general protocol and loaded with compounds AKG-28 and AKG-38 at pH 5.07–5.82 (no buffer substances added) as described in Example 8.

[0357] In an attempt to stabilize liposomes with 0.5 M AS as a sequestering agent against rapid drug release upon contact with blood (as described in Example 17), liposomes using DSPC (generally known to result in stable liposomes with higher drug leakage compared to HSPC) and decreasing percentages of cholesterol (Chol) were prepared and loaded with AKG-28 at a DLO of 250 g / mol PhL or AKG-38 at 600 g / mol PhL (Table 15). Contrary to expectations, reducing the cholesterol content from 40 mol% to 10 mol% cholesterol resulted in a dramatic decrease in encapsulation efficiency for both AKG-28 and AKG-38. Lower cholesterol also destabilized the liposomes against aggregation. Liposomes containing AKG-28 prepared with 1N TEA-SOS and 5 mol% PEG-DSG or PEG-DSPE at 30 mol% cholesterol (70:30 PhL-cholesterol molar ratio) irreversibly aggregated during drug loading, as did formulations containing AKG-38, 30 mol% cholesterol, 5 mol% PEG-DSPE, whereas formulations of AKG-38, 30 mol% cholesterol, 5 mol% PEG-DSG showed reduced loading efficiencies of 77.1% or 462.4 g / mol PhL.

[0358] [Table 21]

[0359] In contrast, liposomes prepared using HSPC and containing 40 mol% or more cholesterol, up to 65 mol% cholesterol (the maximum value tested), showed excellent encapsulation efficiencies of over 87% and no liposome aggregation for both AKG-28 (DL0 250 g / mol PhL) and AKG-38 (DL0 500 g / mol PhL), PEG-lipids (PEG-DSG and PEG-DSPE), and entrapment agents (AS or TEA-SOS) (Table 16).

[0360] Additionally, the ability of the optimized formulations to load linezolid, a current standard of care from this class, was evaluated in both 0.5 M AS and 1 N TEA-SOS formulations. Tedizolid was not sufficiently soluble in water to perform transmembrane gradient-assisted loading into liposomes according to the general protocol in Example 6. In both cases for linezolid, encapsulation efficiency was less than 5%, indicating that these liposomal formulations of AKG-28 and AKG-38 were dramatically superior in their ability to stably encapsulate the drug when compared to linezolid.

[0361] The Z-average size of the liposomes (x z ) and polydispersity index (PDI) were determined by the dynamic light scattering (DLS) cumulant method using a Malvern Zetasizer Pro (Malvern Panalytical) at a measurement angle of 173°.

[0362] [Table 22] JPEG2025501769000099.jpg113159

[0363] [Example 19] In vitro burst release of PEGylated liposomes containing AKG-28 or AKG-38 and varying ratios of phospholipid to cholesterol in the presence of plasma. The in vitro stability of liposomal formulations of AKG-28 and AKG-38 containing 5 mol% PEG-DSPE or PEG-DSG and varying ratios of HSPC to Chol (50–65 mol% Chol) was evaluated in the presence of mouse CD-1 or human pooled plasma (lithium-heparin stabilized from Innovative Research). Plasma was thawed, adjusted to pH 7.4 with 1 N HCl as needed, and filtered sequentially through glass microfiber filters (GF / C), 1 μm polyethersulfone (PES), and 0.22 μm PES filters. Plasma (80 μl) was mixed with the liposomal drug formulation (20 μl) in a 0.5 ml Eppendorf tube. The mixture was then incubated at 37°C for 20 min before being placed in cold water. The mixture (0.1 mL) was chromatographed without delay on a 2 mL Sepharose CL-4B column and eluted with Hepes-buffered saline (pH 7.0), and 0.25 mL of liposomal drug was collected in the void volume fraction. The drug and DiI(3)-DS lipid label were then analyzed by HPLC as described in Example 7, and the remaining encapsulated drug percentage was calculated using the following formula: (A d / A I / (A d,0 / A I,0 ) * 100 = % encapsulated drug remaining (In the formula, A d is that of the drug peak, and A I is the area of ​​the lipid-labeled peak, and A d,0 is the area of ​​the drug peak before incubation with plasma, and A I,0 is that of the lipid-labeled peak before incubation.) was determined using

[0364] The results are shown in Figures 5A, 5B, 5C, and 5D. For liposomes encapsulating AKG-28 (Figure 5A), a burst release phenomenon (a rapid decrease in DL ratio indicating drug release from liposomes) was observed in human plasma for formulations containing 40 mol% cholesterol, but not for formulations containing 45 mol% or more cholesterol. For liposomes encapsulating AKG-38 (Figure 5B), a burst release phenomenon was observed in both human and mouse plasma for formulations containing 40 mol% and 45 mol% cholesterol, but not for formulations containing 50 mol% or more cholesterol.

[0365] [Example 20] In vitro plasma release and in vivo pharmacokinetics of 5 mol% PEG-lipid liposomes containing AKG-38 and 40 or 55 mol% cholesterol. The three liposomal formulations of Example 18 using 0.5M AS scavenger were evaluated in a two-time point pharmacokinetic study in female CD-1 mice as described in Example 7, measuring the percent of injected dose of liposomal lipid remaining in the blood (%ID) at both 5 minutes and 6 hours and measuring drug release from liposomes by determining the drug-to-lipid ratio (DL). Liposomes with 5 mol% PEG-DSG or 5 mol% PEG-DSPE containing 55 mol% Chol showed pre-injection D / L of greater than 95% at 5 minutes and greater than 85% at 6 hours, whereas the PEG-DSG formulation containing 40 mol% Chol showed dramatically reduced DL ratios at both 5 minutes and 6 hours, consistent with the in vitro drug leakage data in the presence of plasma (Table 17). This finding contrasts with previous experience with drug-loaded liposomal formulations, because several highly stable liposomal drugs approved for clinical use, such as PEGylated liposomal doxorubicin and nanoliposomal irinotecan, contain cholesterol at approximately 50 mol% (see, e.g., Doxil® drug information package insert, updated August 2019, and Drummond, DC, et al. (2006). "Development of a highly active nanoliposomal irinotecan using a novel intraliposomal stabilization strategy," Cancer Res. 66(6):3271-3277).

[0366] [Table 23]

[0367] [Example 21] Inhibition of mitochondrial protein synthesis (MPS) by AKG-3, AKG-16, AKG-22, AKG-28, AKG-29, AKG-30, AKG-38, AKG-39, and AKG-40 and selectivity for Mycobacterium tuberculosis (H37Rv) compared with MPS inhibition. Inhibition of mitochondrial protein synthesis was determined using a colorimetric MitoBiogenesis™ intracellular ELISA kit (catalog #ab11021) from AbCam according to the manufacturer's instructions. Mitochondrial protein synthesis inhibition has been correlated with significant toxicities associated with linezolid and other oxazolidinones, most notably ocular and peripheral neuropathy and lactic acidosis (Renslo (2010) Expert Review Anti Infect Ther 8(5)565-574; Flanagan et al. (2015) Antimicrob Agents Chemother 59(1)178-185; Santini et al. (2017) Expert Opin Drug Saf 16(7)833-843). Levels of two mitochondrial proteins were measured simultaneously, including the mitochondrial DNA-encoded subunit I of complex IV (COX-1) and the nuclear DNA-encoded 70 kDa subunit of complex II (SDH-A). The H9C2 rat BDIX cardiomyoblast cell line was used in these studies in a 384-well plate assay format. Cells were grown in DMEM medium containing 10% FBS and 1x glutamine at 37°C and 5% CO2. Cells were seeded at a density of 1,500 cells / well in 384-well plates in 47.5 μl / well. One replicate per concentration and condition of each compound, including nine 3-fold dilutions starting at a high concentration of 200 μM, was added to the cells in 2.5 μl and incubated with the cells for 5 days at 37°C and 5% CO2. Compounds tested included tedizolid and linezolid controls, as well as AKG-3, AKG-16, AKG-22, AKG-28, AKG-29, AKG-30, AKG-38, AKG-39, and AKG-40.

[0368] MitoBiogenesis In-Cell Elisa was then performed according to the manufacturer's instructions (Abcame catalog #ab11021) in a plate reader, with alkaline phosphatase (AP) developed for detection of SDH-1A at 405 nm in a kinetic mode for 15 minutes (20 seconds to 1 minute intervals) and HRP developed for detection of COX-I at 500 nm in a kinetic mode for 15 minutes (20 seconds to 1 minute intervals). COX-I and SDH-A signals were plotted as the ratio of COX-1 / SDH-A against the concentration of each compound, and IC50 values ​​were calculated for each of the nine investigated compounds and two controls.

[0369] The MPS selectivity index (SI-MPS) was determined by dividing the MPS IC in μg / ml by the MIC in the drug-susceptible H37Rv M. tuberculosis strain, determined as described in Example 2. The two compounds tested, AKG-28 and AKG-29, had SI-MPS values ​​more than 10-fold higher than those determined for linezolid and more than 20-fold higher than those determined for tedizolid. Both of these compounds contained a primary amino group at the R2 position of the oxazolidinone ring. Because of its high potency (MIC<0.1) and high selectivity for M. tuberculosis compared with mitochondrial protein synthesis, AKG-28 is an excellent candidate for encapsulation in liposomes and treatment of tuberculosis or other mycobacterial diseases.

[0370] [Table 24]

[0371] [Example 22] [Scale-up preparation of liposomal AKG-28 Lot 275] Lot 267. The general procedure of Example 6 was followed. 4.95 g (6.30 mmol) of HSPC (Lipoid AG), 2.98 g (7.71 mmol) of cholesterol (Dishman, High purity), and 850 mg (0.315 mmol) of PEG-DSPE (Lipoid AG) (molar ratio of HSPC:Chol:PEG-DSPE 45:55:2.25) were mixed with 9 ml of absolute ethanol (Sigma, E-7023) and heated with stirring on a 68°C bath until all lipids were dissolved. In a separate container, 93.3 g of 0.5 M aqueous ammonium sulfate (0.2 μm filtered) was preheated on a 68°C bath and poured into the hot lipid ethanol solution with stirring. The resulting suspension was stirred for 20 minutes in a 68°C bath and then extruded eight times at 260-300 psi through a stack of two 47 mm 100 nm pore-size and one 200 nm pore-size polycarbonate track-etched membranes (Whatman Nucleopore) using a Lipex 100 ml thermobarrel liposome extrusion apparatus (Northern Lipids, Inc.) heated with circulating water at 68°C. The resulting extruded liposomes were kept overnight in a refrigerator (2-8°C) and then filtered under positive pressure through a 0.2 μm polyethersulfone (PES) filter. The extraliposomal scavenger (ammonium sulfate) was removed by TFF buffer exchange into endotoxin-free water on a KrosFlo TFF system using polysulfone hollow fiber cartridges with a MW cutoff of 500 KDa (Spectrum Laboratories) until the residual conductivity fell to less than 200 μS / cm (143 μS / cm after a 5.2-fold volume exchange). The phospholipid concentration in the post-TFF liposome suspension was determined to be 57.4 mM by the phosphomolybdenum blue method.

[0372] 720 mg of AKG-28 (as the dihydrochloride salt) in the form of a 20 mg / ml aqueous stock solution (adjusted to pH 5.03 with NaOH) was mixed with the post-TFF liposome suspension to form a loading mixture with a drug-to-phospholipid (DL) ratio of 250 g / mol in the presence of 45 mg / ml dextrose and 6 mg / ml AKG-28. The mixture was rapidly heated to 60-63 °C by external heating with constant stirring, and the incubation continued with stirring in a 65 °C bath. After a 20-minute incubation, the mixture was rapidly cooled to below 10 °C in ice water and held at this temperature for approximately 10 minutes. After reaching ambient temperature and adjusting to 0.1 M NaCl, the drug-loaded liposomes were purified by TFF using a polysulfone hollow fiber cartridge with a molecular weight cutoff of 500 kD. Liposomes were pre-concentrated to approximately 12 mg / ml AKG-28 by diafiltration and purified from extraliposomal drug by TFF exchange in 10 mM HEPES-Na buffer, pH 7.0, containing 0.144 M NaCl, made with endotoxin-free water (HBS-7 buffer) over a total of eight volume exchanges. The percentage of unencapsulated drug before purification was estimated spectrophotometrically at 305 nm in the pre-concentrated diafiltrate and found to be approximately 0.9% (corresponding to a loading efficiency of 99.1%). The concentrated, purified liposomes were aseptically passed through a 0.2 μm sterile filter and analyzed for particle size by DLS and for drug and phospholipid concentrations by spectroscopy. This procedure was repeated three more times (Lots 269, 271, and 273). The resulting liposomes had the characteristics shown in Table 19.

[0373] [Table 25]

[0374] These lots were mixed to obtain Lot 275, which had 12.0 mg / ml of AKG-28 in liposomal form, particle size Xz 113.7 nm, and PDI 0.0417.

[0375] [Example 23] [Scale-up preparation of liposomal AKG-38 Lot 276] Lot 268. The protocol of Example 22 was used with the following differences: a stock aqueous solution of AKG-38 (as free base) was prepared by dissolving the drug in an equal volume of 1N HCl and adjusting the volume to obtain 20 mg / ml AKG-38 (as free base), pH 5.08. The loading mixture contained 1300 mg of AKG-38, prepared at a DL ratio of 8 mg / ml AKG-38 and 450 g / mol phospholipid, and further contained 10 mM NaCl. The loaded liposomes were pre-concentrated to approximately 22 mg / ml of drug; the percentage of unencapsulated drug before purification was estimated spectrophotometrically at 305 nm in the diafiltrate before concentration and found to be approximately 3.2% (corresponding to a loading efficiency of 96.8%). This process was repeated three more times (Lots 270, 272, and 274). The resulting liposomes had the characteristics shown in Table 20.

[0376] [Table 26]

[0377] These lots were mixed to obtain Lot 276, which had 22.3 mg / ml AKG-38 in liposomal form, particle size Xz 113.1 nm, and PDI 0.0454.

[0378] [Example 24] [Preparation of "empty liposomes" Lot 277] 2 mmol of HSPC, 2.444 mmol of cholesterol, and 0.1 mmol of PEG-DSPE (45:55:2.25 molar ratio of HSPC:Chol:PEG-DSPE) were dissolved in ethanol to form a liposome suspension, which was then extruded through a polycarbonate membrane as described in Example 22, except that the non-exchangeable cation sulfate, 0.13 M sodium sulfate, was used instead of 0.5 M ammonium sulfate. The extruded liposomes were purified from extraliposomal sodium sulfate and loaded into HBS-7 buffer by TFF buffer exchange using a polysulfone hollow fiber cartridge with a MWCO of 500 kDa over a total of 10-fold exchange. The purified liposomes had 42.9 mM phospholipid, a particle size of Xz 113.7 nm, and a PDI of 0.0612. They were aseptically passed through a 0.2 μm sterile filter and adjusted to 20 mM phospholipid with sterile HBS-7.

[0379] [Example 25] [Liposomal AKG-38 Lot 279] The general procedure of Example 6 was followed. 13.102 g (16.67 mmol) of HSPC (Lipoid AG), 7.877 g (20.37 mmol) of cholesterol (Dishman, High purity), and 2.250 g (0.833 mmol) of PEG-DSPE (Lipoid AG) (molar ratio of HSPC:Chol:PEG-DSPE 45:55:2.25) were mixed with 25 ml of absolute ethanol (Sigma, E-7023) and heated with stirring on a 68 °C bath until all lipids were dissolved. In a separate container, 259.1 g (250 ml) of 0.5 M aqueous ammonium sulfate (0.2 μm filtered) was preheated on a 70 °C bath and poured into the hot lipid ethanol solution with stirring. The resulting suspension was stirred on a 70 °C bath for at least 20 minutes and divided into four portions. Each portion was extruded five times at 280 psi through a stack of two 47 mm 100 nm pore-size and one 200 nm pore-size polycarbonate track-etched membranes (Whatman Nucleopore) using a Lipex 100 ml thermobarrel liposome extrusion apparatus (Northern Lipids, Inc.) heated with circulating water at 70°C. These partially extruded liposome portions were mixed (Xz 129.7 nm) and extruded together five more times through the same membrane stack, yielding liposomes with a size of Xz 115.9 nm and a PDI of 0.0212. The liposomes were kept overnight in a refrigerator (2–8°C) and then filtered through a 0.2 μm polyethersulfone (PES) filter under positive pressure. The phospholipid concentration was found to be 60.22 ± 0.34 mM. The extraliposomal scavenger (ammonium sulfate) was removed by TFF buffer exchange into endotoxin-free water on a KrosFlo TFF system using polysulfone hollow fiber cartridges with a MW cutoff of 500 kDa (Spectrum Laboratories) until the residual conductivity dropped to 180 μS / cm after a 5.1-volume exchange. The phospholipid concentration in the post-TFF liposome suspension was determined by the phosphomolybdenum blue method to be 54.97 ± 0.32 mM.

[0380] AKG-38 (free base) was mixed with 0.95 equivalents of 1N HCl and made up with endotoxin-free water to obtain a 20 mg / ml aqueous stock solution (pH 5.16). The solution was passed through a 0.2 μm filter, and an amount of the filtrate containing 3958 mg of drug was mixed with the post-TFF liposome suspension to form a loading mixture with a drug-to-phospholipid (DL) ratio of 450 g / mol in the presence of 44.5 mg / ml dextrose, 10 mM NaCl, and 8 mg / ml AKG-38 at pH 5.54. The mixture was heated to 61°C by external heating with constant stirring for 5 minutes, and incubation continued in a 65°C bath for an additional 22 minutes with stirring. The mixture was then transferred to an ice-water bath, stirred for 7 minutes, the temperature was lowered to 10°C, and the mixture was kept in the ice-water bath for an additional 8 minutes. After removal from the ice bath, allowing to reach ambient temperature, and adjusting to 0.1 M NaCl by adding 3 M NaCl stock, the drug-loaded liposomes (pH 6.53) were purified by TFF using a polysulfone hollow fiber cartridge with a molecular weight cutoff of 500 kD. The liposomes were pre-concentrated to approximately 22 mg / ml AKG-38 by diafiltration and purified from extraliposomal drug by TFF exchange in HBS-7 buffer over a total of eight volume exchanges. The concentrated, purified liposomes were aseptically passed through a 0.2 μm PES high-flow sterile filter and analyzed for particle size by DLS and for drug and phospholipid concentrations by spectroscopy. The liposomes had the following characteristics: AKG-38 21.1 ± 0.19 mg / ml, DL ratio 454 ± 4.7 g / mol phospholipid, Xz 116.4 nm, and PDI 0.0231. The yield of the formulated drug was 3834 mg (96.9%).

[0381] [Example 26] [Liposome AKG-28 Lot 281] The general procedure of Example 6 was followed. Extruded liposomes composed of HSPC, cholesterol, and PEG-DSPE in a molar ratio of 45:55:2.25 containing 0.5 M ammonium sulfate were prepared as described in Example 25. The extraliposomal sequestering agent (ammonium sulfate) was removed by TFF buffer exchange into endotoxin-free water on a KrosFlo TFF system using a polyethersulfone hollow fiber cartridge with a MW cutoff of 500 KDa (Spectrum Laboratories) until the residual conductivity was reduced to 150 μS / cm (4.1 volume exchanges). The phospholipid concentration in the post-TFF liposome suspension was determined to be 55.4 mM by the phosphomolybdenum blue method.

[0382] 969.5 mg of AKG-28 (as dihydrochloride salt) in the form of a 20 mg / ml aqueous stock solution (adjusted to pH 5.24 with NaOH) was mixed with the post-TFF liposome suspension to form a loading mixture with a drug-to-phospholipid (DL) ratio of 250 g / mol in the presence of 44.5 mg / ml dextrose and 6 mg / ml AKG-28. The mixture was heated to 65.4°C in 2.5 minutes by external heating with constant stirring, and incubation continued with stirring in a 65°C bath. After 20 minutes of incubation, the mixture was cooled to 9.3°C in 2.75 minutes in ice water and held in ice water for approximately 10 minutes. The mixture was then allowed to reach ambient temperature and adjusted to 0.1 M NaCl; pH 6.43. 133.4 g of the loading mixture was subjected to purification by TFF using a polysulfone hollow fiber cartridge with a molecular weight cutoff of 500 kD. Liposomes were pre-concentrated to approximately 12 mg / ml AKG-28 by diafiltration and purified from extraliposomal drug by TFF exchange in HBS-7 buffer over a total of 8.1 volume exchanges. The percentage of unencapsulated drug in the pre-concentrated diafiltrate was estimated spectrophotometrically at 302 nm and found to be approximately 0.7% (corresponding to a loading efficiency of 99.3%). The concentrated, purified liposomes were aseptically passed through a 0.2 μm sterile filter and analyzed for particle size by DLS and for drug and phospholipid concentrations by spectroscopy. The liposomes had the following characteristics: AKG-28 13.26 ± 0.21 mg / ml, DL ratio 258.2 ± 3.7 g / mol phospholipid, Xz 117.3 nm, and PDI 0.0421.

[0383] [Example 27] [Liposome AKG-38 Lot 285] The general procedure of Example 6 was followed. Extruded liposomes composed of HSPC, cholesterol, and PEG-DSPE in a molar ratio of 45:55:2.25 containing 0.5 M ammonium sulfate were prepared essentially as described in Example 25. The extraliposomal sequestering agent (ammonium sulfate) was removed by TFF buffer exchange into endotoxin-free water on a KrosFlo TFF system using a polyethersulfone hollow fiber cartridge with a MW cutoff of 500 KDa (Spectrum Laboratories) until the residual conductivity was reduced to 138 μS / cm (5.6-fold exchange). The phospholipid concentration in the post-TFF liposome suspension was determined to be 53.1 mM by the phosphomolybdenum blue method.

[0384] AKG-38 (free base) was mixed with 0.95 equivalents of 1N HCl and made up with endotoxin-free water to obtain a 19.9 mg / ml aqueous stock solution (pH 5.13). The solution was passed through a 0.2 μm filter, and an amount of the filtrate containing 1400 mg of drug was mixed with the post-TFF liposome suspension to form a loading mixture with a drug-to-phospholipid (DL) ratio of 450 g / mol in the presence of 44.5 mg / ml dextrose, 10 mM NaCl, and 8 mg / ml AKG-38 at pH 5.58. The mixture was heated to 63°C by external heating with constant stirring over 2.25 minutes, and incubation continued in the 65°C bath with stirring for a total of 21 minutes. The mixture was then transferred to an ice-water bath, stirred for 3 minutes, the temperature was lowered to 10.3°C, and the mixture was maintained in the ice-water bath for an additional 7 minutes. After removal from the ice bath, allowing to reach ambient temperature, and adjusting to 0.1 M NaCl by adding 3 M NaCl stock, the drug-loaded liposomes (pH 6.70) were purified by TFF using a polysulfone hollow fiber cartridge with a molecular weight cutoff of 500 KD. The liposomes were pre-concentrated to approximately 22 mg / ml AKG-38 by diafiltration and purified from extraliposomal drug by TFF exchange in HBS-7 buffer over a total of 7.7 volume exchanges. The concentrated, purified liposomes had an AKG-38 concentration of 23.1 mg / ml. The drug concentration was adjusted to 20 mg / ml with HBS-7 buffer, and the liposomes were aseptically passed through a 0.2 μm PES high-flow sterile filter and analyzed for particle size by DLS and for drug and phospholipid concentrations by spectroscopy. The liposomes had the following characteristics: AKG-38 20.35±0.26 mg / ml, DL ratio 437.8±6.5 g / mol phospholipid, Xz 121.1 nm, PDI 0.0200. The yield of formulated drug was 1355 mg (96.8%).

[0385] [Example 28] [Liposome AKG-28 Lot 286] Extruded liposomes (HSPC:Chol:PEG-DSPE 45:55:2.25 molar ratio) containing 0.5 M ammonium sulfate and no extraliposomal entrapped agent were obtained as described in Example 27.

[0386] 600 mg of AKG-28 (as the dihydrochloride salt) in the form of a 20 mg / ml aqueous stock solution (adjusted to pH 5.18 with NaOH) was mixed with the post-TFF liposome suspension to form a loading mixture with a drug-to-phospholipid (DL) ratio of 250 g / mol in the presence of 44.5 mg / ml dextrose and 6 mg / ml AKG-28. The mixture was placed in a 65°C water bath with stirring and allowed to reach 60°C in 4.5 minutes. Incubation with stirring continued for a total of 20 minutes, after which the mixture was cooled to 10.0°C in ice water in 2 minutes and held in the ice-water bath for approximately 10 minutes. The mixture was then allowed to reach ambient temperature and adjusted to 0.1 M NaCl; pH 6.23. 104.6 g of the loading mixture was subjected to purification by TFF using a polysulfone hollow fiber cartri...

Claims

1. Formula (I) 【Chemistry 1】 [In the formula, R １ is a tetrazole ring substituted at the 2' position with an aminoalkyl; R ２ is an amine or acetamide. or a pharma- ceutically acceptable salt thereof, The compound of formula (I) or a pharma- ceutically acceptable salt thereof is encapsulated in liposomes in an aqueous medium having a pH above 6.7; A liposome composition, wherein the liposome comprises phosphatidylcholine, cholesterol, and a PEG polymer-conjugated lipid, and has 50-65 mol % cholesterol relative to the total of cholesterol and non-PEGylated phospholipid in the liposome.

2. R ２ Acetamide (NHCOCH ３ 2. The liposome composition according to claim 1 .

3. R １ but, 【Chemistry 2】 The liposome composition of claim 2 , selected from the group consisting of:

4. The liposome composition of claim 3 , wherein the PEG polymer-conjugated lipid is in an amount of 5 mol % relative to the phosphatidylcholine.

5. 5. The liposomal composition of claim 4, wherein the sulfate salt of the compound of formula (I) is encapsulated in liposomes comprising the phosphatidylcholine, cholesterol and PEG polymer-conjugated lipid in a molar ratio of 45:55:2.

25.

6. The liposome composition of claim 5 , wherein the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soy phosphatidylcholine (HSPC).

7. 7. The liposome composition of claim 6, wherein the PEG polymer-conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE).

8. 8. The liposome composition of claim 7, further comprising a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA, said chelating agent being at a concentration of 0.1-1 mM.

9. The compounds of formula (I) are AKG-38, AKG-39 and AKG-40. 【Table 1】 The liposome composition according to any one of claims 1 to 8, wherein the liposome composition is a compound selected from the group consisting of:

10. 10. The liposome composition of claim 9, wherein the compound of formula (I) is a sulfate salt of AKG-38.

11. The liposome composition of claim 10 , wherein the pH of the liposome composition is greater than 7 and not greater than 8.

12. The liposome composition according to claim 10, wherein the pH of the liposome composition is from 7.3 to 7.

7.

13. The liposome composition of claim 10, wherein the pH of the liposome composition is 7.

5.

14. The compound of formula (I) is AKG-38 【Chemistry 3】 2. The liposome composition of claim 1, wherein the compound is a sulfate salt of said compound, which is encapsulated in liposomes formed from hydrogenated soy phosphatidylcholine (HSPC), cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 in an aqueous medium having a pH of 7.3 to 7.

7.

15. 15. The liposome composition of claim 14, further comprising a chelating agent, said chelating agent being deferoxamine (DFO), said chelating agent being at a concentration of 0.1-1 mM.

16. The liposome composition of claim 14 or 15, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 430-680 g / mol.

17. The liposome composition of claim 14 or 15, wherein the drug / lipid ratio of AKG-38 to total phospholipids (PhL) in the composition is 600 g / mol.

18. a. the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm; b. the liposome composition has a polydispersity index of less than 0.15; The liposome composition according to claim 14 or 15.

19. The liposome composition according to claim 14 or 15, wherein the ratio of encapsulated AKG-38 to total AKG-38 is at least 90%.

20. The liposome composition of claim 14 or 15, wherein the aqueous medium further comprises sodium chloride.

21. a. the aqueous medium has an osmotic pressure of 270 to 330 mOsmol / kg; b. the sodium chloride is at a concentration of 130-150 mM; c. the chelating agent is at a concentration of 0.5 mM; 21. The liposome composition of claim 20.

22. 21. The liposome composition of claim 20, wherein the aqueous medium comprises ammonium ions at a concentration of 20 to 60 mM and the sodium chloride is at a concentration of 50 to 80 mM.

23. 23. The liposome composition of claim 21 or 22, further comprising HEPES or a phosphate buffer.

24. AKG-38 liposome composition having a pH of at least 7.0 and no greater than 8.0, said liposome composition comprising the lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1, and AKG-38 【Chemistry 4】 and a pharma- ceutically acceptable salt thereof. The liposome composition has the following characteristics: a. the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm, or the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 100-130 nm; b. the liposome composition has a polydispersity index of less than 0.15, or the liposome composition has a polydispersity index of less than 0.10; c. The drug / lipid ratio of the AKG-38 in the liposome composition to the total phospholipids (PhL) is 430-480 g / mol, or the drug / lipid ratio of the AKG-38 in the liposome composition to the total phospholipids (PhL) is 500-650 g / mol, or the drug / lipid ratio of the AKG-38 in the liposome composition to the total phospholipids (PhL) is 430-650 g / mol, or the AKG-38 in the liposome composition the drug / lipid ratio of said AKG-38 to total phospholipids (PhL) in said liposome composition is 450 g / mol, or the drug / lipid ratio of said AKG-38 to total phospholipids (PhL) in said liposome composition is 450 g / mol, or the drug / lipid ratio of said AKG-38 to total phospholipids (PhL) in said liposome composition is 600 g / mol, or the drug / lipid ratio of said AKG-38 to total phospholipids (PhL) in said liposome composition is 600 g / mol; d. the total concentration of AKG-38 in said liposomal composition is 12-25 mg / mL, or the total concentration of AKG-38 in said liposomal composition is 13.5-16.5 mg / mL, or the total concentration of AKG-38 in said composition is 15 mg / mL, or the total concentration of AKG-38 in said liposomal composition is 16 mg / mL (160 mg in a 10 mL vial); e) the ratio of encapsulated AKG-38 to the total AKG-38 in said liposome composition is at least 90%, or the ratio of encapsulated AKG-38 to the total AKG-38 in said liposome composition is at least 95%, or the ratio of encapsulated AKG-38 to the total AKG-38 in said liposome composition is at least 97%, or the ratio of encapsulated AKG-38 to the total AKG-38 in said liposome composition is at least 98%; f. the liposome composition comprises an aqueous medium containing sodium chloride and, optionally, ammonium ions; g. the aqueous medium has an osmolality of 270 to 330 mOsmol / kg, or the aqueous medium has an osmolality of 270 to 310 mOsmol / kg; h. the aqueous medium comprises ammonium ions at a concentration of 20-60 mM, or the aqueous medium comprises ammonium ions at a concentration of 50-80 mM, or the aqueous medium comprises ammonium ions at a concentration of less than 0.5 mM, or the aqueous medium comprises ammonium ions at a concentration of less than 1 mM, or the aqueous medium comprises ammonium ions at a concentration of 1-10 mM; i. the aqueous medium comprises sodium chloride at a concentration of 130-150 mM; j. the aqueous medium further comprises a buffer, wherein the buffer buffers the liposome composition at a pH of 7.3 to 7.7 or at a pH of 7.5; k. the aqueous medium further comprises HEPES or a phosphate buffer, or the aqueous medium further comprises HEPES or a phosphate buffer at a concentration of 5-50 mM, or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 20 mM; l. the aqueous medium further comprises a chelating agent, or the aqueous medium further comprises a chelating agent at a concentration of 0.1-1 mM, or the aqueous medium further comprises a chelating agent at a concentration of 0.5 mM, or the aqueous medium further comprises deferoxamine (DFO) or EDTA, or the aqueous medium further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1-1 mM, or the aqueous medium further comprises deferoxamine (DFO) or EDTA at a concentration of 0.5 mM; m. the liposome composition is storage stable; or n. The AKG-38 is encapsulated in the liposome as a sulfate salt of AKG-38. The AKG-38 liposomal composition, further characterized by any one or more of:

25. in an aqueous medium containing a chelating agent selected from the group consisting of deferoxamine (desferrioxamine, desferal), ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), nitrilotriacetic acid (NTA), ethylene glycol-O,O'-bis(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA), N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid (HEDTA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA); Formulated with (5R)-3-{3-fluoro-4-[6-(2-(2-dimethylaminoethyl)-2H-tetrazol-5-yl)-3-pyridinyl]phenyl}-5-(methylacetamido)-1,3-oxazolidin-2-one or a pharma- ceutically acceptable salt thereof; An isotonic AKG-38 liposomal dispersion encapsulated in liposomes containing hydrogenated soy phosphatidylcholine (HSPC), cholesterol, and (PEG(molecular weight 2,000)-distearoylphosphatidylethanolamine, PEG-DSPE) (PEG(2000)-DSPE).

26. 26. The isotonic AKG-38 liposomal dispersion of claim 25 having a pH greater than 6.7 and up to 8.

0.

27. a. the isotonic AKG-38 liposomal dispersion has a pH of 7.5; b. the liposomes are formed from hydrogenated soy phosphatidylcholine (HSPC), cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25, and the chelating agent is deferoxamine; 27. The isotonic AKG-38 liposomal dispersion of claim 26.

28. 27. The isotonic AKG-38 liposomal dispersion of claim 25 or 26, wherein the liposomal dispersion comprises lipid vesicles formed from a composition comprising phosphatidylcholine, 55 mol% cholesterol and 5 mol% PEG-DSG or 5 mol% PEG-DSPE.

29. 29. A method for treating a Methicillin-resistant Staphylococcus aureus (MRSA) bacterial infection, comprising administering to a subject in need thereof a therapeutically effective amount of the liposome composition of any one of claims 1 to 28.

30. 29. A method of making a liposome composition according to any one of claims 1 to 28, comprising preparing an extruded lipid suspension, a. dissolving one or more phospholipids, cholesterol and a PEG-lipid derivative in ethanol to obtain a lipid solution; b. combining the lipid solution of step (a) with a sequestering agent solution to obtain a homogenous lipid suspension having a desired phospholipid concentration; c. extruding the lipid suspension of step (b) through a membrane with defined pore size, such as a polycarbonate track-etched (PCTE) membrane with a nominal pore size of 50-200 nm; d. purifying liposomes from extraliposomal entrapped agents in the extruded lipid suspension to obtain a purified extruded liposome preparation; e. contacting the liposomes with the compound of formula (I) in an aqueous medium to effect encapsulation of the compound in the liposomes; f. Optionally, removing unencapsulated compounds; g. providing said liposomes in a physiologically acceptable medium suitable for parenteral use; Including, the scavenger solution of step (b) comprises an aqueous solution of ammonium sulfate at a concentration greater than 0.25 M; The method of claim 1, wherein the physiologically acceptable medium of step (g) comprises a chelating agent.

31. 31. The method of claim 30, wherein the capture agent solution in step (b) comprises ammonium sulfate at a concentration of 0.5 M.

32. 31. The method of claim 30, wherein the chelating agent is deferoxamine.

33. 31. The method of claim 30, wherein the extruded liposomes in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm.

34. R ２ However, amine (NH ２ 2. The liposome composition of claim 1 .

35. R １ but, 【Chemistry 5】 35. The liposome composition of claim 34, selected from the group consisting of:

36. The liposome composition of claim 34, wherein the PEG polymer-conjugated lipid is in an amount of 5 mol % relative to the phosphatidylcholine.

37. a. the sulfate salt of the compound of formula (I) is encapsulated in liposomes comprising the phosphatidylcholine, cholesterol and PEG polymer-conjugated lipids in a molar ratio of 45:55:2.25; b. the phosphatidylcholine is distearoylphosphatidylcholine (DSPC) or hydrogenated soy phosphatidylcholine (HSPC); c. The PEG polymer conjugated lipid is PEG (molecular weight 2,000)-distearoylglycerol (PEG-DSG) or PEG (molecular weight 2,000)-distearoylphosphatidylethanolamine (PEG-DSPE); 37. The liposome composition of claim 36.

38. 38. The liposome composition of any one of claims 34 to 37, further comprising a chelating agent selected from the group consisting of deferoxamine (DFO) and EDTA, said chelating agent being at a concentration of 0.1 to 1 mM.

39. The compound of formula (I) is AKG-28, AKG-29, AKG-30, AKG-31, AKG-38 and AKG-39. 【Table 2】 The liposome composition of claim 38, wherein the compound is selected from the group consisting of:

40. The compound of formula (I) is AKG-28 【Chemistry 6】 40. The liposome composition of claim 38, wherein the compound is a sulfate salt of said compound, which is encapsulated in liposomes formed from hydrogenated soy phosphatidylcholine (HSPC), cholesterol and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 in an aqueous medium having a pH of 7.3 to 7.

7.

41. AKG-28 liposome composition comprising liposomes, the liposomes comprising the lipids HSPC, cholesterol, and PEG(2000)-DSPE in a molar ratio of 45:55:2.25 or a mass ratio of 5:3:1, and AKG-28 encapsulated in the liposomes. 【Chemistry 7】 and a pharma- ceutically acceptable salt thereof. The liposome composition has the following characteristics: a. the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm, or the liposome composition comprises mono- or oligolamellar vesicles having a z-average diameter of 100-130 nm; b. the liposome composition has a polydispersity index of less than 0.15, or the liposome composition has a polydispersity index of less than 0.10; c. The drug / lipid ratio of the AKG-28 to the total phospholipids (PhL) in the composition is 99-530 g / mol PhL, or 85-456 g / mol as AKG-28 free base (FB); 99-470 g / mol PhL, or 85-400 g / mol as AKG-28 free base (FB); 230-280 g / mol, or 190-240 g / mol as FB; or the drug / lipid ratio of the AKG-28 to the total phospholipids (PhL) in the composition is 290-360 g / mol, or 245-305 g / mol as FB; or the drug / lipid ratio of the AKG-28 to the total phospholipids (PhL) in the composition is 300-340 g / mol, or 256-2 or the drug / lipid ratio of said AKG-28 to total phospholipids (PhL) in said composition is 250 g / mol, or 215 g / mol as FB; or the drug / lipid ratio of said AKG-28 to total phospholipids (PhL) in said composition is 330 g / mol, or 280 g / mol as FB; or the drug / lipid ratio of said AKG-28 to total phospholipids (PhL) in said composition is 330 g / mol, or 280 g / mol as FB; d. the total concentration of AKG-28 in said composition is 8-15 mg / mL, or 6.8-12.8 mg / mL as FB; or the total concentration of AKG-28 in said composition is 9-11 mg / mL, or 7.6-9.4 mg / mL as FB; or the total concentration of AKG-28 in said composition is 10 mg / mL, or 8.5 mg / mL as FB; or the total concentration of AKG-28 in said composition is 10 mg / mL (100 mg in a 10 ml vial), or 8.5 mg / mL as FB (85 mg in a 10 ml vial); e. the ratio of encapsulated AKG-28 to the total AKG-28 in said liposome composition is at least 90%;or the ratio of encapsulated AKG-28 to the total AKG-28 in said liposome composition is at least 95%;or the ratio of encapsulated AKG-28 to the total AKG-28 in said liposome composition is at least 97%;or the ratio of encapsulated AKG-28 to the total AKG-28 in said liposome composition is at least 98%; f. the liposome composition comprises an aqueous medium containing sodium chloride and, optionally, ammonium ions; g. the aqueous medium has an osmolarity of 270-330 mOsmol / kg, or the aqueous medium has an osmolarity of 270-310 mOsmol / kg, or the aqueous medium is isotonic; h. the aqueous medium comprises ammonium ions at a concentration of 20-60 mM, or the aqueous medium comprises ammonium ions at a concentration of 50-80 mM, or the aqueous medium comprises ammonium ions at a concentration of less than 0.5 mM, or the aqueous medium comprises ammonium ions at a concentration of less than 1 mM, or the aqueous medium comprises ammonium ions at a concentration of 1-10 mM, or the aqueous medium comprises ammonium ions at a concentration of less than 130 mM; i. the aqueous medium further comprises a buffer, wherein the buffer buffers the liposome composition at a pH of 7.3 to 7.7 or at a pH of 7.5; j. the aqueous medium further comprises HEPES or a phosphate buffer, or the aqueous medium further comprises HEPES or a phosphate buffer at a concentration of 5-50 mM, or the aqueous medium comprises HEPES or a phosphate buffer at a concentration of 20 mM; k. the composition further comprises a chelating agent, or the composition further comprises a chelating agent at a concentration of 0.1-1 mM, or the composition further comprises a chelating agent at a concentration of 0.5 mM, or the composition further comprises deferoxamine (DFO) or EDTA, or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of 0.1-1 mM, or the composition further comprises deferoxamine (DFO) or EDTA at a concentration of 0.5 mM, or l. AKG-28 is encapsulated in liposomes as the sulfate salt of AKG-28. The AKG-28 liposomal composition, further characterized by any one or more of:

42. 42. A method of making a liposome composition according to any one of claims 34 to 41, comprising preparing an extruded lipid suspension, a. dissolving one or more phospholipids, cholesterol and a PEG-lipid derivative in ethanol to obtain a lipid solution; b. combining the lipid solution of step (a) with a sequestering agent solution to obtain a homogenous lipid suspension having a desired phospholipid concentration; c. extruding the lipid suspension of step (b) through a membrane with defined pore size, such as a polycarbonate track-etched (PCTE) membrane with a nominal pore size of 50-200 nm; d. purifying liposomes from extraliposomal entrapped agents in the extruded lipid suspension to obtain a purified extruded liposome preparation; e. contacting the liposomes with the compound of formula (I) in an aqueous medium to effect encapsulation of the compound in the liposomes; f. Optionally, removing unencapsulated compounds; g. providing said liposomes in a physiologically acceptable medium suitable for parenteral use; Including, the scavenger solution of step (b) comprises an aqueous solution of ammonium sulfate at a concentration greater than 0.25 M; The method of claim 1, wherein the physiologically acceptable medium of step (g) comprises a chelating agent.

43. 43. The method of claim 42, wherein the capture agent solution of step (b) comprises ammonium sulfate at a concentration of 0.5 M.

44. 43. The method of claim 42, wherein the chelating agent is deferoxamine.

45. 43. The method of claim 42, wherein the extruded liposomes in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm.

46. 44. The method of claim 43, wherein the chelating agent is deferoxamine and the liposomes extruded in step (c) are mono- or oligolamellar vesicles having a z-average diameter of 90-130 nm.

47. 42. A method for treating a mycobacterial infection, comprising administering to a subject in need thereof a therapeutically effective amount of the liposome composition of any one of claims 34-41.

48. 48. The method of claim 47, wherein the mycobacterial infection is an infection with Mycobacterium tuberculosis, or an infection with a multidrug resistant (MDR) strain of Mycobacterium tuberculosis, or an infection with an extremely drug resistant (XDR) strain of Mycobacterium tuberculosis.

49. Use of the liposomal composition according to any one of claims 1 to 28 for treating a methicillin-resistant Staphylococcus aureus (MRSA) bacterial infection.

50. Use of the liposomal composition according to any one of claims 34 to 41 for treating a mycobacterial infection.

51. 29. The liposomal composition of any one of claims 1 to 28 for use in a method for treating a methicillin-resistant Staphylococcus aureus (MRSA) bacterial infection comprising administering to a subject in need thereof a therapeutically effective amount of the liposomal composition.

52. 42. The liposomal composition of any one of claims 34 to 41 for use in a method of treating a mycobacterial infection comprising administering to a subject in need thereof a therapeutically effective amount of the liposomal composition.

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