Laquinimod is a compound which has been shown to be effective in the acute experimental autoimmune encephalomyelitis (aEAE) model (U.S. Pat. No. 6,077,851). Its chemical name is N-ethyl-N-phenyl-1,2,-dihydro-4-hydroxy-5-chloro-1-methyl-2-oxoquinoline-3-carboxamide, and its Chemical Registry number is 248281-84-7.
Laquinimod is a small molecule having the following chemical structure:
Laquinimod sodium has high oral bioavailability and has been suggested as an oral formulation for the treatment of Multiple Sclerosis (MS). (Polman, C. et al., (2005) “Treatment with laquinimod reduces development of active MRI lesions in relapsing MS”, Neurology. 64:987-991; Sandberg-Wollheim M, et al. (2005) “48-week open safety study with high-dose oral laquinimod in patients”, Mult Scler. 11:S154). Studies have also shown that laquinimod can reduce development of active MRI lesions in relapsing MS. (Polman, C. et al., (2005) “Treatment with laquinimod reduces development of active MRI lesions in relapsing MS”, Neurology. 64:987-991).
In order to prepare laquinimod as a pharmaceutical drug product, manufacturing processes need to take into consideration the possibility of the impurities disclosed herein being present in the product. Early syntheses of laquinimod produced product contaminated with impurities such as methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate and 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline (K. Jansson et al., (2006) “Synthesis and Reactivity of Laquinimod, a Quinoline-3-carboxamide: Intramolecular Transfer of the Enol Proton to a Nitrogen Atom as a Plausible Mechanism for Ketene Formation, J. Org. Chem., 71, 1658-1667). Several other impurities have also been disclosed (U.S. Pat. No. 6,077,851; U.S. Pat. No. 6,875,869; and U.S. Pat. No. 7,884,208). U.S. Patent Application Publication No. 2012/0010239, the contents of which are incorporated by reference into this application, discloses the impurity 5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. U.S. Patent Application Publication No. 2013/0217724, the contents of which are incorporated by reference into this application, discloses the impurity N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. U.S. Patent Application Publication No. 2013/00345256, the contents of which are incorporated by reference into this application, discloses the impurity N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. U.S. Pat. No. 8,178,127, the contents of which are incorporated by reference into this application, discloses the impurities 2-chloro-6-(1-ethyl-N-methyl-2-oxoindoline-3-carboxamido)benzoic acid, 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[1]ethylindolin-[2]-one and 5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl,1,2,3,4-tetrahydroquinoline-3-carboxamide.
The processes of synthesis of laquinimod and the preparation of its sodium salt are disclosed in U.S. Pat. No. 6,077,851. An additional process of synthesis of laquinimod acid, but not laquinimod sodium, having low levels of certain impurities is disclosed in U.S. Pat. No. 6,875,869. U.S. Pat. No. 7,884,208 teaches a process for the preparation of laquinimod sodium which removes certain impurities present after the salt formation step, thus resulting in a crystalline mixture of higher purity as well as a crystalline mixture having large particles, and improved tapped and bulk density. Pharmaceutical compositions comprising laquinimod sodium are disclosed in PCT International Application Publication No. WO 2005/074899.
In the preparation of laquinimod sodium disclosed in U.S. Pat. No. 6,077,851, laquinimod acid was so suspended in ethanol, and 5M sodium hydroxide solution was added. After stirring, the resulting precipitate was filtered, washed with ethanol, and dried. The method used to make laquinimod sodium in U.S. Pat. No. 6,077,851 is commonly referred to as a slurry-to-slurry salt formation.
In the slurry-to-slurry salt formation method of U.S. Pat. No. 6,077,851, the laquinimod sodium is not dissolved in solution. Any solid impurities, if present in the laquinimod sodium suspension, are therefore not removed by filtration.
U.S. Pat. No. 6,875,869 discloses a process of preparing the base compound laquinimod in high yield and low level of impurities. However, the process in U.S. Pat. No. 6,875,869 is only for synthesis of the base compound (laquinimod acid) and not the salt. As such, the slurry-to-slurry salt formation process would still be needed to form the sodium salt.
U.S. Pat. No. 7,884,208 teaches an improved process for preparing laquinimod sodium resulting in crystals of higher purity as well as crystals having improved crystalline characteristics, e.g., comprising no more than 2 ppm of a heavy metal and having higher tapped density. In the processes disclosed in Examples 13-17 of U.S. Pat. No. 7,884,208, laquinimod sodium is dissolved in water to form an aqueous solution; the aqueous solution is concentrated; and then a water-miscible anti-solvent is added to the concentrated solution to form laquinimod sodium crystals. The process of U.S. Pat. No. 7,884,208 removes the impurities after salt formation, thus resulting in laquinimod sodium of higher purity than the laquinimod sodium produced directly from the “slurry to slurry” process of U.S. Pat. No. 6,077,851.
The subject invention provides a mixture of crystalline laquinimod sodium particles, wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and wherein:
The subject invention provides a mixture of crystalline laquinimod sodium particles, wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 40 microns, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 15 microns, and (iii) 10% or more of the total amount by volume of the laquinimod sodium panicles have a size of less than 5 microns and wherein:
The subject invention provides a pharmaceutical composition comprising the mixture as described herein and a pharmaceutically acceptable carrier.
The subject invention provides a process of recrystallization of laquinimod sodium comprising:
The subject invention also provides for a mixture of crystalline laquinimod sodium particles prepared by the process as described herein, and a pharmaceutical composition comprising said mixture.
The subject invention provides a mixture of crystalline laquinimod sodium particles, wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have size of 15 microns or less, and wherein
The subject invention also provides a pharmaceutical composition comprising the mixture as described herein and a pharmaceutically acceptable carrier.
The subject invention also provides an isolated compound having the structure:
or a salt thereof.
The subject invention also provides an composition comprising a compound having the structure:
The subject invention also provides a pharmaceutical composition comprising an amount of laquinimod and at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE), wherein a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or f) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
The subject invention also provides a process for preparing 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) comprising the steps of: a) adding sodium hydroxide solution to a suspension of 5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide in water, b) stirring the mixture of step a) followed by addition of hydrochloric acid solution, c) extracting the aqueous solution with ethyl acetate, d) washing the organic phase with brine, e) drying the organic phase over sodium sulfate, f) filtering the suspension, g) evaporating the filtrate, h) purifying the residue by crystallization from isopropyl alcohol, i) cooling the suspension followed by filtering and washing with isopropyl alcohol, and j) obtaining and drying the resulting white solid.
The subject invention also provides 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-1-3-HLAQ) prepared by the process described above.
The subject invention also provides a process for preparing 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) comprising the steps of: a) heating a mixture of 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) and dimethylsulfoxide, b) cooling the mixture of step a), and c) filtering the mixture of step b) and collecting the resulting filtrate.
The subject invention also provides 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) prepared by the process described above.
The subject invention also provides a process for preparing 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) comprising the steps of: a) heating a mixture of ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in a solution of hydrochloric acid in acetic acid, b) cooling the mixture of step a), c) diluting the mixture of step b) with 2-propanol and further cooling the diluted mixture, and d) filtering off the crystals resulting from step c).
The subject invention also provides 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) prepared by the process described above.
The subject invention also provides a process for preparing Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) comprising the steps of: a) forming sodium dimethylmalonate by reaction of dimethylmalonate in dimethlformamide with sodium methoxide solution, b) reacting the intermediate 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione with sodium dimethylmalonate to form methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) sodium salt, and c) acidifying methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) sodium salt to methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME).
The subject invention also provides Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) prepared by the process described above.
The subject invention also provides a process for preparing Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) comprising the steps of: a) adding sodium hydride to a solution of 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione and diethyl malonate in dimethylformamide, b) heating the mixture of step a) while stirring, c) cooling the solution of step b), d) quenching the reaction mixture of step c), e) acidifying the mixture of step d), f) filtering then drying the mixture of step e), and g) crystallizing the crude product of step f) by dissolving in ethanol following by slow cooling.
The subject invention also provides Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) prepared by the process described above.
The subject invention also provides a process for testing whether a sample of laquinimod contains an undesirable impurity which comprises determining whether the sample contains a compound having the structure:
The subject invention also provides a process for preparing a validated pharmaceutical composition comprising laquinimod comprising: a) obtaining a batch of laquinimod; b) determining the amount of at least one of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the batch using by an HPLC method; and c) preparing the pharmaceutical composition from the batch only if
The subject invention also provides a process for preparing a pharmaceutical composition comprising laquinimod, or for distributing a validated batch of a pharmaceutical composition comprising laquinimod, comprising a) obtaining a batch of laquinimod or of the pharmaceutical composition; b) performing stability testing with a sample of the batch; c) determining the total amount of at least one of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sample of the batch after stability testing by an HPLC method; and d) validating the batch for distribution or preparing the pharmaceutical composition from the batch only if the sample of the batch after stability testing contains
The subject invention also provides a process for validating a batch of a pharmaceutical product containing laquinimod or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier for distribution comprising a) subjecting a sample of the batch to stability testing; b) determining the amount of at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-1-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sample of the batch after stability testing by an HPLC method; and c) validating the batch for distribution only if the sample of the batch after stability testing contains
The subject invention also provides a process for preparing a packaged pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof comprising: a) obtaining a batch of pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof; b) performing stability testing with a sample from the batch; c) determining the amount of at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sample by an HPLC method after stability testing; and d) packaging the pharmaceutical composition in only if
The subject invention also provides an impurity or a salt thereof for use, as a reference standard to detect trace amounts of the impurity in a pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof, wherein the impurity is selected from the group consisting of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a diluent solution comprising the buffer solution and acetonitrile, f) Preparing a blank solution comprising the diluent solution and aqueous acetonitrile, g) Injecting into the HPLC the resolution solution, the blank solution, the standard solution, and the sample solution, h) Running the HPLC using ultraviolet absorption at 240 nm and the diluent solution as the mobile phase, i) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and j) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2 dihydroquinoline-3-carboxamide (5-HLAQ) or Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a blank solution comprising the buffer solution and acetonitrile, f) Injecting into the HPLC the resolution solution, the blank solution, the standard solution, and the sample solution, g) Running the HPLC using ultraviolet absorption at 240 nm and a mobile phase of a mixture of the buffer solution, and acetonitrile, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA), 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[1]ethylindolin-[2]-one] (SPIRO-LAQ), or 5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide (3-HLAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a Quantitation Limit (QL) solution comprising the impurity, d) Preparing a resolution solution comprising laquinimod and the impurity, e) Preparing a buffer solution by dissolving ammonium dihydrogen phosphate in water, and adjusting to pH of 7.0±0.10 with aqueous ammonia or phosphoric acid, f) Preparing a blank solution comprising the buffer solution and acetonitrile, g) Injecting into the HPLC the resolution solution, the blank solution, the QL solution, the standard solution and the sample solution, h) Running the HPLC using a ultraviolet absorption at 212 nm and a mobile phase of a mixture of the buffer solution, acetonitrile, and methanol, i) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and j) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) or N-ethyl-4-hydroxy-1-methyl-5-(2,3,4,5,6-pentahydroxyhexylamino)-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to PH or 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a blank solution comprising the buffer solution and acetonitrile, f) Injecting into the HPLC the resolution solution, the blank solution, the standard solution and the sample solution, g) Running the HPLC using a ultraviolet absorption at 242 nm and the blank solution as the mobile phase, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) or N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2 dihydroquinoline-3-carboxamide (5-HLAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a blank solution comprising methanol and acetonitrile, e) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, f) Injecting into the HPLC the resolution solution, the blank solution, and the sample solution, g) Running the HPLC using a ultraviolet absorption at 240 nm and a mobile phase comprising acetonitrile and the buffer solution, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA), 5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide (3-HLAQ) or 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[1]ethylindolin-[2]-one] (SPIRO-LAQ).
Laquinimod is a small molecule having the following chemical structure:
It is an oral immunomodulator which has demonstrated therapeutic effect in various experimental inflammatory/autoimmune animal models, such as Experimental Autoimmune Encephalomyelitis (EAE), an animal model for Multiple Sclerosis (MS), Dextran Sodium Solphate (DSS) induced colitis for Inflammatory Bowel Disease, Non-Obese Diabetic (NOD) mice for Type I Diabetes (IDDM), Experimental Autoimmune Neuritis (EAN) for Guillain-Barre Syndrome, Systemic Lupus Erythematosus (SLE), lupus nephritis, lupus arthritis, Crohn's Disease and Rheumatoid arthritis. The therapeutic activity of laquinimod in these models results from a variety of mechanistic effects, including reduction of leukocyte infiltration into target tissues by modulation of chemokine-mediated T-cell adhesion, modulation of cytokine balance, down regulation of MHC class II resulting in alteration of antigen presentation, and effects on dendritic cells subpopulations (PCT International Application Publication No. WO2013/169746).
A pharmaceutically acceptable salt of laquinimod includes lithium, sodium, potassium, magnesium, calcium, manganese, copper, zinc, aluminum and iron. Salt formulations of laquinimod and the process for preparing the same are described, e.g., in U.S. Patent Application Publication No. 2005/0192315 and PCT International Application Publication No. WO 2005/074899, which are hereby incorporated by reference into this application.
The subject invention provides a mixture of crystalline laquinimod sodium particles, wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and wherein:
In an embodiment of the mixture, (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 40 microns or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 15 microns.
In an embodiment of the mixture, 10% or more of the total amount by volume of the laquinimod sodium particles have a size of 5 microns or less and wherein:
The subject invention provides a mixture of crystalline laquinimod sodium particles, wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 40 microns, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 15 microns, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 5 microns and wherein:
In an embodiment of the mixture, the mixture is prepared in a single batch comprising 2.5 kg or more of laquinimod sodium. In an embodiment of the mixture, the laquinimod sodium particles are determined based on an unmilled sample of the mixture. In an embodiment of the mixture, the size and amount by volume of laquinimod sodium particles are determined based on a milled sample of the mixture.
In an embodiment of the mixture, the mixture has a bulk density of 0.2 g/mL to 0.4 mL. In an embodiment of the mixture, the mixture has a tapped density of 0.40 g/mL to 0.7 g/mL.
In an embodiment of the mixture, an amount of aluminium in the mixture is less than 5 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of aluminium in the mixture is less than 2 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of calcium in the mixture is less than 60 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of calcium in the mixture is less than 25 ppm relative to the amount by weight of laquinimod sodium, hi an embodiment of the mixture, an amount of copper in the mixture is less than 1 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of copper in the mixture is less than 0.6 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of zinc in the mixture is less than 7 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of zinc in the mixture is less than 4 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of heavy metal in the mixture is no more than 20 ppm relative to the amount by weight of laquinimod sodium.
In an embodiment of the mixture, a total amount of polar impurities in the mixture is no more than 1.00% relative to the amount of laquinimod sodium as measured by HPLC.
In an embodiment of the mixture, an amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture is no more than 0.15% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) in the mixture is no more than 0.15% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) in the mixture is no more than 0.12% relative to the amount of laquinimod sodium as measured by HPLC. In another embodiment, the amount of MCQME in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, a total amount of non-polar impurities in the mixture is no more than 0.50% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of N-ethylaniline (NEA) in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (5-HLAQ) in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of 5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (DELAQ) in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC.
In an embodiment of the mixture, an amount of laquinimod acid in the mixture is no more than 1.00% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of dimethyl malonate in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of diethyl malonate in the mixture is no more than 0.10% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the mixture, an amount of dimethyl sulfate in the mixture is no more than 1 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of water in the mixture is no more than 1.5% by weight relative to the amount of laquinimod sodium as measured by K.F. coulometric titration. In an embodiment of the mixture, an amount of sodium from 5.8% to 6.4% relative to the amount by weight of laquinimod sodium.
In an embodiment of the mixture, an amount of ethanol in the mixture is no more than 5000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of n-heptane in the mixture is no more than 5000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of n-octane in the mixture is no more than 2000 ppm relative to the amount by weight of laquinimod sodium. In another embodiment of the mixture, an amount of n-octane in the mixture is no more than 200 ppm relative to the amount by weight of laquinimod sodium. In another embodiment of the mixture, an amount of n-octane in the mixture is no more than 20 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of methanol in the mixture is no more than 3000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of acetone in the mixture is no more than 5000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of dioxane in the mixture is no more than 380 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the mixture, an amount of dimethyl formamide in the mixture is no more than 880 ppm relative to the amount by weight of laquinimod sodium.
In an embodiment of the mixture, the recited compound is present in the mixture. In another embodiment, the recited compound is present in at least trace amounts.
The subject invention provides a pharmaceutical composition comprising the mixture of the subject invention and a pharmaceutically acceptable carrier.
In an embodiment of the pharmaceutical composition, a total amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) in the pharmaceutical composition is no more than 0.50% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the pharmaceutical composition, a total amount of polar impurities in the pharmaceutical composition is no more than 2.00% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the pharmaceutical composition, an amount of N-ethylaniline (NEA) in the pharmaceutical composition is no more than 0.50% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the pharmaceutical composition, an amount of 5-chloro-N-ethyl-3-hydroxy-1-methyl-5-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide (3-HLAQ) in the pharmaceutical composition is no more than 0.50% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the pharmaceutical composition, a total amount of non-polar impurities in the pharmaceutical composition is no more than 1.00% relative to the amount of laquinimod sodium as measured by HPLC. In an embodiment of the pharmaceutical composition, an amount of N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ) in the pharmaceutical composition is no more than 1.00% relative to the amount of laquinimod sodium as measured by HPLC.
In an embodiment of the pharmaceutical composition, an amount of water in the pharmaceutical composition is no more than 1.50% relative to the amount of laquinimod sodium as measured by K.F. coulometric titration. In an embodiment of the pharmaceutical composition, an amount of water in the pharmaceutical composition is no more than 0.80% relative to the amount of laquinimod sodium as measured by K.F. coulometric titration. In an embodiment of the pharmaceutical composition, an amount of sodium from 5.8% to 6.4% relative to the amount by weight of laquinimod sodium.
In an embodiment of the pharmaceutical composition, an amount of ethanol in the pharmaceutical composition is no more than 5000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of n-heptane in the pharmaceutical composition is no more than 5000 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of n-octane in the pharmaceutical composition is no more than 2000 ppm relative to the amount by weight of laquinimod sodium. In another embodiment of the pharmaceutical composition, an amount of n-octane in the pharmaceutical composition is no more than 200 ppm relative to the amount by weight of laquinimod sodium. In another embodiment of the pharmaceutical composition, an amount of n-octane in the pharmaceutical composition is no more than 20 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of methanol in the pharmaceutical composition is no more than 380 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of acetone in the pharmaceutical composition is no more than 880 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of dioxane in the pharmaceutical composition is no more than 380 ppm relative to the amount by weight of laquinimod sodium. In an embodiment of the pharmaceutical composition, an amount of dimethyl formamide in the pharmaceutical composition is no more than 880 ppm relative to the amount by weight of laquinimod sodium.
In an embodiment of the pharmaceutical composition, the recited compound is present in the pharmaceutical composition. In another embodiment, the recited compound is present in at least trace amounts.
The subject invention provides a method of treating a subject afflicted with a form of multiple sclerosis, lupus nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, a GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated disorder, or an ocular inflammatory disorder comprising administering to the subject the mixture of the subject invention or the pharmaceutical composition of the subject invention so as to thereby treat the subject.
The subject invention provides a method for alleviating a symptom of multiple sclerosis, lupus nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, a GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated disorder, or an ocular inflammatory disorder in a subject afflicted with a form of multiple sclerosis comprising administering to the subject the mixture of any one of the subject invention or the pharmaceutical composition of the subject invention thereby alleviating the symptom of multiple sclerosis in the subject.
In one embodiment, the mixture or the pharmaceutical composition for use in the treatment of, or alleviation of symptoms of, a form of multiple sclerosis, lupus nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, a GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated disorder, or an ocular inflammatory disorder.
The subject invention provides a use of the mixture or the pharmaceutical composition for the manufacture of a medicament for treating, or alleviating a symptom of, a form of multiple sclerosis, a GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated disorder, lupus nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, or an ocular inflammatory disorder.
The subject invention provides a process of recrystallization of laquinimod sodium comprising: a) dissolving an amount of laquinimod sodium in water to form an aqueous solution; b) concentrating the aqueous solution to form a concentrated solution comprising approximately 1.7-1.8 mL of water per gram of laquinimod sodium; c) adding acetone to the concentrated solution of step b); and d) isolating recrystallized laquinimod sodium.
In an embodiment of the process, the amount of laquinimod sodium in step a) is 2.5 kg or greater. In an embodiment of the process, step a) is performed with 10-12 mL of water per gram of laquinimod sodium. In an embodiment of the process, step a) is performed with approximately 11 mL of water per gram of laquinimod sodium. In an embodiment of the process, step a) is performed by heating the aqueous solution to a temperature of 58-75° C. In an embodiment of the process, step a) is performed by heating the aqueous solution to a temperature of 60-73° C.
In an embodiment of the process, crystallization occurs after step a) and before step c).
In an embodiment of the process, crystallization is induced by rapid stirring during or after the concentrating step b). In an embodiment of the process, crystallization is induced by addition of a seed crystal during or after the concentrating step b). In an embodiment of the process, crystallization occurs without addition of a seed crystal.
In an embodiment of the process, step b) is performed under conditions appropriate to induce crystallization at the concentration of 1.7-1.8 mL of water per gram of laquinimod sodium. In an embodiment of the process, step b) is performed at 28-45° C. hi an embodiment of the process, step b) is performed at 30-40° C. In an embodiment of the process, step c) is performed with the concentrated solution at 40-55° C. In an embodiment of the process, step c) is performed with the concentrated solution at 45-50° C. In an embodiment of the process, step c) is performed with 6-12 mL of acetone per gram of laquinimod sodium. In an embodiment of the process, step c) is performed with approximately 10 mL of acetone per gram of laquinimod sodium. In an embodiment of the process, step c) is performed over a period of 1-4 hours. In an embodiment of the process, step c) is performed over a period of 1.2-2.5 hours. In an embodiment of the process, step c) is followed by cooling the solution to a temperature no less than −14° C. and no more than 6° C. In an embodiment of the process, step c) is followed by cooling the solution to a temperature no less than −4° C. and no more than 4° C.
In an embodiment of the process, the solution is cooled over a period of 3-5 hours. In an embodiment of the process, the solution is cooled over a period of 3.5-4.5 hours.
In an embodiment of the process, step d) further comprises washing the recrystallized laquinimod sodium with 1-4 mL of acetone per gram of crude laquinimod sodium used in step a). In an embodiment of the process, step d) further comprises washing the recrystallized laquinimod sodium with approximately 3 mL of acetone per gram of crude laquinimod sodium used in step a). In an embodiment of the process, step d) further comprises drying the recrystallized laquinimod sodium for no less than one hour at 30-40° C. under a vacuum of no more than 50 mmHg.
In an embodiment of the process, the isolated recrystallized laquinimod sodium in step d) is a mixture of crystalline laquinimod sodium particles having a particle size distribution such that (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 5 microns or less.
The subject invention provides a mixture of crystalline laquinimod sodium particles prepared by the process of the subject invention.
In an embodiment of the mixture of crystalline laquinimod sodium particles prepared by the process of the subject invention, (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of 5 microns or less.
In an embodiment of the mixture,
In an embodiment of the mixture,
In one embodiment, the amount of n-octane in the mixture is no more than 200 ppm. In another embodiment, the amount of n-octane in the mixture is no more than 20 ppm.
In an embodiment of the mixture, the recited compound is present in the mixture. In another embodiment, the recited compound is present in at least trace amounts.
The subject invention provides a pharmaceutical composition comprising the mixture of crystalline laquinimod sodium particles prepared by the process of the subject invention, and a pharmaceutically acceptable carrier.
In an embodiment of the pharmaceutical composition,
In an embodiment of the pharmaceutical composition,
In one embodiment, the amount of n-octane in the pharmaceutical composition is no more than 200 ppm relative to the amount by weight of laquinimod sodium. In another embodiment, the amount of n-octane in the pharmaceutical composition is no more than 20 ppm relative to the amount by weight of laquinimod sodium.
The subject invention provides a mixture of crystalline laquinimod sodium particles wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and wherein a) an amount of aluminium in the mixture is less than 5 ppm relative to the amount by weight of laquinimod sodium; b) an amount of calcium in the mixture is less than 60 ppm relative to the amount by weight of laquinimod sodium; c) an amount of copper in the mixture is less than 1 ppm relative to the amount by weight of laquinimod sodium; or d) an amount of zinc in the mixture is less than 7 ppm relative to the amount by weight of laquinimod sodium.
In an embodiment of the pharmaceutical composition, the recited compound is present in the pharmaceutical composition. In another embodiment, the recited compound is present in at least trace amounts.
The subject invention provides a pharmaceutical composition comprising the mixture of the subject invention and a pharmaceutically acceptable carrier.
In an embodiment of the mixture, 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 1 micron or greater, 2 microns or greater, 3 microns or greater or 4 microns or greater. In an embodiment of the mixture, 90% of the total amount by volume of the laquinimod sodium particles have a size of greater than 1 micron, greater than 2 microns, greater than 3 microns, greater than 4 microns or greater than 5 microns.
In a further embodiment of the mixture, the mixture has a tapped density of 0.4 g/mL to 0.7 g/mL, 0.44 g/mL to 0.7 g/mL, 0.45 g/mL to 0.7 g/mL, 0.46 g/mL to 0.7 g/mL or 0.5 g/mL to 0.7 g/mL. In a further embodiment of the mixture, the mixture has a bulk density of 0.2 g/mL to 0.4 g/mL, 0.2 g/mL to 0.33 g/mL, 0.2 g/mL to 0.31 g/mL.
In a further embodiment of the pharmaceutical composition, the pharmaceutical composition comprises mannitol. In a further embodiment of the pharmaceutical composition, the pharmaceutical composition comprises meglumine. In a further embodiment of the pharmaceutical composition, the pharmaceutical composition comprises sodium stearyl fumarate.
In a further embodiment of the pharmaceutical composition, not less than 70% of the labeled amount of laquinimod is dissolved in 30 minutes. In a further embodiment of the pharmaceutical composition, capsules of the pharmaceutical composition contain 90-110% of the labeled amount of laquinimod. In a further embodiment of the pharmaceutical composition, capsules of the pharmaceutical composition contain 95-105% of the labeled amount of laquinimod. In a further embodiment of the pharmaceutical composition, capsules of the pharmaceutical composition contain 98.0-102.0% of the labeled amount of laquinimod.
In a further embodiment of the pharmaceutical composition, the pharmaceutical composition has content uniformity conforming to the U.S. Pharmacopeia. In a further embodiment of the pharmaceutical composition, the pharmaceutical composition has content uniformity conforming to European Pharmacopeia.
The subject invention also provides an isolated compound having the structure:
or a salt thereof.
The subject invention also provides a composition comprising a compound having the structure:
The subject invention also provides a pharmaceutical composition comprising an amount of laquinimod and at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE), wherein a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or f) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount not more than 10%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.12%, relative to the concentration of laquinimod, based on a determination by an HPLC method. In another embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an less than 0.06%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount greater than 0.02%, and not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.02%, and not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount greater than 0.02%, and not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.02%, and not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount greater than 0.02%, and not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.12%, relative to the concentration of laquinimod, based on a determination by an HPLC method. In another embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method. In yet another embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount greater than 0.05%, and not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.05%, or greater than 0.10%, and not more than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount greater than 0.05%, and less than 0.15%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.06%, and not more than 0.10%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount greater than 0.05%, and not more than 0.10%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.12%, relative to the concentration of laquinimod, based on a determination by an HPLC method. In another embodiment, methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.02%, or greater than 0.03%, and less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.02%, and less than 0.06%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, the composition further comprises at least one pharmaceutically acceptable carrier.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount not more than 1.0%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) the combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) the combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an less than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or f) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or g) the combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is less than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.02%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount greater than 0.02%, and not more than 1.0%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.03%, and not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) the combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is greater than 0.03%, and not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, N-Ethylaniline (NEA) is present in the pharmaceutical composition in an amount not more than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount greater than 0.05%, and not more than 1.0%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.1%, and not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) the combined amount of 5-Chloro-4-hydroxy-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is greater than 0.1%, and not more than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In an embodiment of the pharmaceutical composition as described herein, a) 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or c) 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.5%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or d) Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical composition in an greater than 0.03%, and less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or f) Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) is present in the pharmaceutical composition in an amount greater than 0.02%, and less than 0.05%, relative to the concentration of laquinimod, based on a determination by an HPLC method, or g) the combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) present in the pharmaceutical composition is greater than 0.03%, and less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount greater than 0.03%, relative to the concentration of laquinimod, based on a determination by an HPLC method. In another embodiment, 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is present in the pharmaceutical composition in an amount less than 0.1%, relative to the concentration of laquinimod, based on a determination by an HPLC method.
In one embodiment, the pharmaceutical composition as described herein further comprises laquinimod sodium salt. In another embodiment, the pharmaceutical composition is in an oral unit dosage form. In another embodiments, it is in the form of a capsule, a tablet, or a liquid suspension. In another embodiment, the oral unit dosage form comprises more than 0.3 mg laquinimod. In another embodiments, the oral unit dosage form comprises more than 0.5 mg laquinimod. In yet another embodiment, the oral unit dosage form comprises more than 0.6 mg laquinimod.
The subject invention also provides a process for preparing 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) comprising the steps of: a) adding sodium hydroxide solution to a suspension of 5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide in water, b) stirring the mixture of step a) followed by addition of hydrochloric acid solution, c) extracting the aqueous solution with ethyl acetate, d) washing the organic phase with brine, e) drying the organic phase over sodium sulfate, f) filtering the suspension, g) evaporating the filtrate, h) purifying the residue by crystallization from isopropyl alcohol, i) cooling the suspension followed by filtering and washing with isopropyl alcohol, and j) obtaining and drying the resulting white solid.
The subject invention also provides 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) prepared by the process described above.
The subject invention also provides a process for preparing 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) comprising the steps of: a) heating a mixture of 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) and dimethylsulfoxide, b) cooling the mixture of step a), and c) filtering the mixture of step b) and collecting the resulting filtrate.
The subject invention also provides 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) prepared by the process described above.
The subject invention also provides a process for preparing 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) comprising the steps of: a) heating a mixture of ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in a solution of hydrochloric acid in acetic acid, b) cooling the mixture of step a), c) diluting the mixture of step b) with 2-propanol and further cooling the diluted mixture, and d) filtering off the crystals resulting from step c).
The subject invention also provides 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) prepared by the process described above.
The subject invention also provides a process for preparing Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) comprising the steps of: a) forming sodium dimethylmalonate by reaction of dimethylmalonate in dimethlformamide with sodium methoxide solution, b) reacting the intermediate 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione with sodium dimethylmalonate to form methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) sodium salt, and c) acidifying methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) sodium salt to methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME).
The subject invention also provides Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) prepared by the process described above.
The subject invention also provides a process for preparing Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) comprising the steps of: a) adding sodium hydride to a solution of 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione and diethyl malonate in dimethylformamide, b) heating the mixture of step a) while stirring, c) cooling the solution of step b), d) quenching the reaction mixture of step c), e) acidifying the mixture of step d), f) filtering then drying the mixture of step e), and g) crystallizing the crude product of step f) by dissolving in ethanol following by slow cooling.
The subject invention also provides Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) prepared by the process described above.
The subject invention also provides a process for testing whether a sample of laquinimod contains an undesirable impurity which comprises determining whether the sample contains a compound having the structure:
The subject invention also provides a process for preparing a validated pharmaceutical composition comprising laquinimod comprising: a) obtaining a batch of laquinimod; b) determining the amount of at least one of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the batch using by an HPLC method; and c) preparing the pharmaceutical composition from the batch only if
In one embodiment, in step c) the pharmaceutical composition is prepared from the batch only if the batch is determined to have not more than 0.12% Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), relative to the concentration of laquinimod. In another embodiment, in step c) the pharmaceutical composition is prepared from the batch only if the batch is determined to have not more than 0.1% Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), relative to the concentration of laquinimod.
The subject invention also provides a process for preparing a pharmaceutical composition comprising laquinimod, or for distributing a validated batch of a pharmaceutical composition comprising laquinimod, comprising a) obtaining a batch of laquinimod or of the pharmaceutical composition; b) performing stability testing with a sample of the batch; c) determining the total amount of at least one of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sample of the batch after stability testing by an HPLC method; and d) validating the batch for distribution or preparing the pharmaceutical composition from the batch only if the sample of the batch after stability testing contains
In one embodiment, the process further comprises step e) distributing the batch if in step d) the batch is validated for distribution. In another embodiments, in step d) the batch is validated for distribution or the pharmaceutical composition is prepared from the batch only if the sample of the batch after stability testing contains not more than 0.12% Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) relative to the concentration of laquinimod. In another embodiment, in step d) the batch is validated for distribution or the pharmaceutical composition is prepared from the batch only if the sample of the batch after stability testing contains not more than 0.1% Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), relative to the concentration of laquinimod.
The subject invention also provides a process for validating a batch of a pharmaceutical product containing laquinimod or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier for distribution comprising a) subjecting a sample of the batch to stability testing; b) determining the amount of at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sale of the batch after stability testing by an HPLC method; and c) validating the batch for distribution only if the sample of the batch after stability testing contains
In an embodiment, in step c) the batch is validated for distribution only if the sample of the batch after stability testing not more than a total of the 0.1% of N-Ethylaniline (NEA) relative to the concentration of laquinimod.
The subject invention also provides a process for preparing a packaged pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof comprising; a) a) obtaining a batch of pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof; b) performing stability testing with a sample from the batch; c) determining the amount of at least one of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) in the sample by an HPLC method after stability testing; and d) d) packaging the pharmaceutical composition in only if
In one embodiment, in step d) the pharmaceutical composition is packaged only if the content of N-Ethylaniline (NEA) in the sample is determined to be not more than 0.1% to the concentration of laquinimod.
In one embodiments, the laquinimod is laquinimod sodium salt.
The subject invention also provides an impurity or a salt thereof for use, as a reference standard to detect trace amounts of the impurity in a pharmaceutical composition comprising laquinimod or a pharmaceutically acceptable salt thereof, wherein the impurity is selected from the group consisting of 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ), 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), N-Ethylaniline (NEA), and Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a diluent solution comprising the buffer solution and acetonitrile, f) Preparing a blank solution comprising the diluent solution and aqueous acetonitrile, g) Injecting into the HPLC the resolution solution, the blank solution, the standard solution, and the sample solution, h) Running the HPLC using ultraviolet absorption at 240 nm and the diluent solution as the mobile phase, i) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and j) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME). N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2 dihydroquinoline-3-carboxamide (5-HLAQ) or Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a blank solution comprising the buffer solution and acetonitrile, f) Injecting into the HPLC the resolution solution, the blank solution, the standard solution, and the sample solution, g) Running the HPLC using ultraviolet absorption at 240 nm and a mobile phase of a mixture of the buffer solution, and acetonitrile, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA), 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[l]ethylindolin-[2]-one](SPIRO-LAQ), or 5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide (3-HLAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a Quantitation Limit (QL) solution comprising the impurity, d) Preparing a resolution solution comprising laquinimod and the impurity, e) Preparing a buffer solution by dissolving ammonium dihydrogen phosphate in water, and adjusting to pH of 7.0±0.10 with aqueous ammonia or phosphoric acid, f) Preparing a blank solution comprising the buffer solution and acetonitrile, g) Injecting into the HPLC the resolution solution, the blank solution, the QL solution, the standard solution and the sample solution, h) Running the HPLC using a ultraviolet absorption at 212 nm and a mobile phase of a mixture of the buffer solution, acetonitrile, and methanol, i) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and j) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid (BH-3-HLAQ) or N-ethyl-4-hydroxy-1-methyl-5-(2,3,4,5,6-pentahydroxyhexylamino)-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to PH or 7.0±0.05 with aqueous ammonia or glacial acetic acid, e) Preparing a blank solution comprising the buffer solution and acetonitrile, f) Injecting into the HPLC the resolution solution, the blank solution, the standard solution and the sample solution, g) Running the HPLC using a ultraviolet absorption at 242 nm and the blank solution as the mobile phase, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA), Methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME) or N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2 dihydroquinoline-3-carboxamide (5-HLAQ).
The subject invention also provides a method of determining the concentration of an impurity in a pharmaceutical composition comprising laquinimod and a pharmaceutically acceptable carrier, the method comprising, a) Preparing a sample solution from the pharmaceutical composition, b) Preparing a standard solution comprising the impurity, c) Preparing a resolution solution comprising laquinimod and the impurity, d) Preparing a blank solution comprising methanol and acetonitrile, e) Preparing a buffer solution by dissolving ammonium acetate in water, and adjusting to pH of 7.0±0.05 with aqueous ammonia or glacial acetic acid, f) Injecting into the HPLC the resolution solution, the blank solution, and the sample solution, g) Running the HPLC using a ultraviolet absorption at 240 nm and a mobile phase comprising acetonitrile and the buffer solution, h) Determining the retention time (RT) and the areas of the peaks of the impurity in the chromatograms of the sample solution, and i) Performing quantitation of the impurity with respect to the corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA), 5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide (3-HLAQ) or 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[l]ethylindolin-[2]-one](SPIRO-LAQ).
Each embodiment disclosed herein is contemplated as being applicable to each of the other disclosed embodiments. Thus, all combinations of the various elements described herein are within the scope of the invention. For example, any embodiment for an element of a mixture a contemplated as being applicable to a pharmaceutical composition. As another example, an embodiment to a tapped density of 0.46 g/mL to 0.7 g/mL is contemplated as being applicable to a mixture having an amount of aluminium in the mixture less than 5 ppm relative to the amount by weight of laquinimod sodium.
Applicants have unexpectedly achieved a mixture of crystalline laquinimod sodium that is improved over the laquinimod sodium mixture of the prior art.
U.S. Pat. No. 7,884,208 teaches a process for recrystallizing laquinimod sodium which produces a mixture of crystalline laquinimod sodium having larger crystals, lowered impurity levels and certain improved crystalline characteristics as compared to the laquinimod sodium crystals known at the time.
Specifically, the process of U.S. Pat. No. 7,884,208 achieves a mixture of crystalline laquinimod sodium having (i) a mixture wherein 10% or more of the total amount by volume of the laquinimod sodium particles has a size of greater than 40 microns and wherein 50% or more of the total amount by volume of the laquinimod sodium particles has a size of greater than 15 microns, (ii) high density (tapped and bulk), (iii) low heavy metal content, and (iv) low content of certain polar impurities.
However, the recrystallization process of U.S. Pat. No. 7,884,208 (Examples 13-17, Tables 1-4) does not produce a mixture of recystallized laquinimod sodium particles wherein 90% or more of the total amount by volume of the laquinimod sodium have a size of 40 microns or less, or 50% or more of the total amount by volume of the laquinimod sodium has a size of 15 microns or less.
Likewise, the recrystallization process of U.S. Pat. No. 7,884,208 does not produce a mixture of recystallized laquinimod sodium particles wherein 90% or more of the total amount by volume of the laquinimod sodium have a size of less than 40 microns, wherein 50% or more of the total amount by volume of the laquinimod sodium has a size of less than 15 microns, and wherein 10% or more of the total amount by volume of the laquinimod sodium has a size of less than 5 microns.
Example 14 of U.S. Pat. No. 7,884,208 produced a mixture of recystallized laquinimod sodium particles wherein 10% or more of the total amount by volume of the laquinimod sodium have a size of less than 5 microns. However, this Example also shows reduced quality of crystalline characteristics, specifically Tapped Density. The mixture produced by Example 14 has an acceptable D(0.1) value but an undesired Tapped Density.
Conversely, Example 13 of U.S. Pat. No. 7,884,208 produced a mixture having high Tapped Density, but did not produce crystals wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less.
Importantly, U.S. Pat. No. 7,884,208 is unable to achieve the advantages of recrystallization, i.e., better density and impurity profiles, while also producing laquinimod sodium crystals wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, or 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less.
U.S. Pat. No. 7,884,208, by way of Example 1 (batches A, B and C) and Tables 1-3, also teaches that the process disclosed in U.S. Pat. No. 6,077,851 results in a mixture of crystalline laquinimod sodium having all four of (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, or 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, (ii) poor density (tapped and bulk), (iii) high heavy metal content, and (iv) high polar impurity content. Importantly, although U.S. Pat. No. 6,077,851 achieves a mixture wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or, 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, but it does not achieve crystals having acceptable density or low levels of impurities.
The prior art contains no teaching of a process for preparing laquinimod sodium wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, or 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and having desirable density and purity profiles.
The present invention provides a process which is capable of producing a mixture of recystallized laquinimod sodium crystals wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less or (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and having desirable density and purity profiles, specifically, (i) high density (tapped or bulk), (ii) low heavy metal content, or (iii) low polar impurity content.
The mixture of laquinimod sodium provided by the present invention is achieved by an improved recrystallization process.
The laquinimod sodium manufactured by the recrystallization processes of the present invention has improved purity and density profiles over the laquinimod sodium disclosed in U.S. Pat. No. 6,077,851 and improved crystalline characteristics, especially smaller particles, over U.S. Pat. No. 7,884,208.
The modified recrystallization process of the present invention unexpectedly results in different recrystallization conditions than achieved by the process disclosed in U.S. Pat. No. 7,884,208 and, thusly, results in different products. Specifically, concentrating the aqueous solution to 1.7-1.8 unexpectedly results in crystalline laquinimod sodium particles having reduced levels of impurities, improved crystalline characteristics, and wherein 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and 10% or more of the total amount by volume of the laquinimod sodium particles have a size of 5 microns or less.
Without being limited to any one particular theory, an important factor affecting crystallization is initial concentration of crystallizing solution. Careful control of concentration of the solution can facilitate initiation of spontaneous crystallization prior to the end of the concentration step. The process of the present invention can initiate spontaneous crystallization prior to addition of acetone.
Concentrating the aqueous solution to form a concentrated solution comprising approximately 1.7-1.8 ml of water per gram of laquinimod sodium is an important aspect of the present invention.
U.S. Pat. No. 7,884,208 teaches advantages associated with large particles. Specifically, U.S. Pat. No. 7,884,208 teaches that larger particles of laquinimod sodium are more “processable” when making pharmaceutical compositions and that smaller particles are often associated with dust-like properties which may interfere with processing, and sometimes associated with flowability problems which may interfere with manufacturing. Further, U.S. Pat. No. 7,884,208 teaches that chemical stability has been shown to be decreased by the increase in surface area that results from smaller particle size. (Felmeister, A. Chpt 88, Remington's Pharmaceutical Sciences, 15th Edition, Mack Publishing Company, Easton, Pa. (1975)).
However, the subject invention has unexpectedly achieved an improved mixture of laquinimod sodium wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of 5 microns or less.
Laquinimod has been graded as a highly potent API, demanding special caution and avoiding material contact with workers and environment. Laquinimod has been graded as having the highest potency rate, corresponding to a recommended acceptable daily intake (ADI) during operations and manufacturing of less than 0.01 mg/day or <1 μg/m3 as an 8-hour TWA. High potency compounds are associated with controls, whether engineering, administrative or procedure-related, that afford the desired level of worker protection. For example, high potency compounds may require no human intervention or manual operations. (Bruce D. Naumann, Control Banding In The Pharmaceutical Industry, http://www.aioh.org.au/downloads/documents/ControlBandingBNaumaun.pdf)
Importantly, although the mixture of the subject invention may be milled or unmilled, the present invention is achieved without the need of a milling operation.
The mixture of laquinimod sodium of the present invention overcomes the potential problems associated with mixtures having large particles. With respect to processing and manufacturing, the small size of the laquinimod sodium particles of the present invention may obviate the need for milling and comminution steps. U.S. Pat. No. 7,884,208 reflected the understanding in the art that there are problems associated with small particles in pharmaceutical compositions, but the laquinimod sodium of the present invention has no problems associated with stability, processing or manufacturing.
Comminution introduces its own set of problems to a drug substance beyond the disadvantages of introducing an additional drug processing step. For example, milling can introduce impurities, new polymorphs, amorphous sections in the crystalline structure of the API, other changes to particle morphology, differences in agglomeration, increased solubility, changes in moisture levels, and changes in compressibility (Hausner, “The Role of Particle Size in the Development of Generic Products” 2003). As a result, comminution may affect the efficacy and safety of a drug substance. Some of the disadvantages of comminution are illustrated by the side effects resulting from microcrystalline Nitrofurantoin compared to macrocrystalline Nitrofurantoin. (Brumfitt, W. and J. M. T. Hamilton-Miller, J. Antimicrobial Chemotherapy 42:363:371 (1998)).
Accordingly, it is advantageous to produce a drug substance which does not contain large particles to avoid the inefficiencies of additional process steps such as milling or sieving. The laquinimod sodium of the present invention provides a mixture of laquinimod sodium particles having small particle sizes which avoids safety problems and additional problems related to milling.
Another concern during formulation processes is maintaining uniformity of content of the drug product. In the case of laquinimod, the unit dose of laquinimod is quite low relative to the total weight of the drug product, e.g., tablet or capsule. A typical formulation, for example, may comprise only a small amount of laquinimod, e.g., 0.3, 0.6 or 1.2 mg, in a capsule with total weight of over 200 mg. As such, small fluctuation in the amount of laquinimod due to problems of flowability, segregation, uniformity, or poor homogenous distribution could result in a large percent deviation from the desired amount, e.g., 0.3, 0.6 or 1.2 mg. The mixture of laquinimod sodium of the present invention provides high uniformity of content and minimal fluctuations in the amount of laquinimod in the capsules.
Uniformity of the shape of laquinimod particles is also an important concern during formulation as a lack of uniformity of shape can cause variation in density of drug substance and cause problems during drug product formation, e.g., capsule or tablet formation. Crystalline laquinimod sodium particles are rod-shaped particles. It is known that milling operations may result in changes to particle shape.
Decreased particle size is known to result in faster dissolution profiles. The rate of dissolution of small particles is usually faster than that of large particles because a greater surface area of the drug substance is in contact with the liquid medium. When formulating a drug with a low dissolution rate, it is desirable to decrease particle size in order to increase dissolution and thus facilitate rapid gastrointestinal or oral absorption.
In such cases where drug substances have no recognized problems associated with dissolution rate, particle size reduction may be inadvisable and even deleterious. Increasing surface area can increase degradation rates of the drug substance. As discussed, for example, in U.S. Pat. Nos. 8,178,127 and 7,989,473, laquinimod sodium is susceptible to degradation.
Unexpectedly, in spite of known disadvantages associated with small particle sizes, it was found that an improved drug substance and drug product resulted from a mixture of crystalline laquinimod sodium particles wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of 40 microns or less, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of 15 microns or less, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of 5 microns or less.
Accordingly, an advantage of the recrystallization process of the present invention is that the resulting mixture of crystalline laquinimod sodium has particles having small particle sizes, which is associated with high uniformity and homogeneity with respect to distribution of the API into capsules, tablets and other drug products. Laquinimod sodium crystals having small particle sizes can obviate or reduce the need for additional milling steps. The small particle sizes of the laquinimod sodium of the present invention are achieved without sacrificing desirable purity or density profiles and without the need for prior milling operations.
Another advantage of the present invention is that laquinimod sodium crystals have a higher density than the laquinimod sodium crystals produced by the slurry-to-slurry process of U.S. Pat. No. 6,077,851. Low tapped density is anathema to certain prized qualities in a drug substance or drug product such as compressibility, the ability of a powder to decrease in volume under pressure, and compactability, the ability of a powder to be compressed into a tablet of certain strength or hardness. Crystals with low tapped density are also known to have poor flowability, which results in a lack of uniformity of content in finished dosage forms, especially in tablets. (Rudnic et al. Chpt. 45, Remington's Pharmaceutical Sciences, 20th Edition, Lippincott Williams & Wilkins, Baltimore, Md. (2000)) Uniformity of content is especially important for pharmaceutical compositions comprising a potent drug substance, e.g., Laquinimod sodium.
Compared to the slurry-to-slurry process of U.S. Pat. No. 6,077,851, the present invention also shows low aggregation of the particles and, additionally, provides particles with acceptable density and lower levels of impurities. As shown in
Another advantage of the present invention is that the process of the present invention is environmentally friendly without sacrificing desirable crystalline characteristics. Specifically, by use of water as the primary solvent, the present invention achieves both environmental friendliness and improved crystalline characteristics, specifically with respect to particle size distribution over U.S. Pat. No. 7,884,208.
As used herein, and unless stated otherwise, each of the following terms shall have the definition set forth below.
As used herein, “laquinimod” means laquinimod acid or a pharmaceutically acceptable salt thereof, including laquinimod sodium.
As used herein, “laquinimod acid” is N-ethyl-N-phenyl-1,2,-dihydro-4-hydroxy-5-chloro-1-methyl-2-oxoquinoline-3-carboxamide, and its Chemical Registry number is 248281-84-7. “Laquinimod sodium” is the sodium salt of laquinimod acid.
As used herein, D(0.1) is the particle size, in microns, below which 10% by volume distribution of the population is found.
As used herein, D(0.5) is the particle size, in microns, below which 50% by volume distribution of the population is found.
As used herein, D(0.9) is the particle size, in microns, below which 90% by volume distribution of the population is found.
As used herein, “crystalline characteristics” includes particle size distribution, bulk density and tapped density.
As used herein, “drug substance” refers to the active ingredient in a drug product or for use in a drug product, which provides pharmacological activity or other direct effect in the diagnosis, cure, mitigation, treatment, or prevention of disease, or to affect the structure or any function of the body of man or animals.
As used herein, “drug product” refers to the formulated or finished dosage form containing the drug substance as well as at least one pharmaceutically acceptable carrier.
As used herein, a composition that is “free” of a chemical entity means that the composition contains, if at all, an amount of the chemical entity which cannot be avoided following an affirmative act intended to purify the composition by separating the chemical entity from the composition.
For example, a composition which is “free” of an alkalizing agent means that the alkalizing agent, if present at all, is a minority component of the composition by weight. Preferably, when a composition is “free” of a component, the composition comprises less than 0.1 wt %, 0.05 wt %, 0.02 wt %, or 0.01 wt % of the component.
As used herein, an “isolated” compound is a compound isolated from the crude reaction mixture following an affirmative act of isolation. The act of isolation involves separating the compound from the other known components of the crude reaction mixture, with some impurities, unknown side products and residual amounts of the other known components of the crude reaction mixture permitted to remain. Purification is an example of an affirmative act of isolation.
As used herein, “stability testing” refers to tests conducted at specific time intervals and various environmental conditions (e.g., temperature and humidity) to see if and to what extent a drug product degrades over its designated shelf life time. The specific conditions and time of the tests are such that they accelerate the conditions the drug product is expected to encounter over its shelf life. For example, detailed requirements of stability testing for finished pharmaceuticals are codified in 21 C.F.R §211.166, the entire content of which is hereby incorporated by reference.
As used herein, “dissolution rate” is determined based on the amount of drug substance dissolved in 30 min. as indicated in the U.S. Pharmacopeia <711>.
As used herein, “atmospheric pressure” refers to a pressure of about 1 atm.
As used herein, “ambient temperature” refers to a temperature of about 20° C. to about 30° C.
As used herein, “about” in the context of a numerical value or range means ±10% of the numerical value or range recited or claimed.
As used herein, “approximately” in the context of a numerical value or range means ±5% of the numerical value or range recited or claimed.
The term “stable pharmaceutical composition” as used herein in connection with the composition according to the invention denotes a composition, which preserves the physical stability/integrity and/or chemical stability/integrity of the active pharmaceutical ingredient during storage.
Furthermore, “stable pharmaceutical composition” is characterized by its level of degradation products not exceeding 5% at 40° C./75% RH after 6 months or 3% at 55° C./75% RH after two weeks, compared to their level in time zero.
As used herein, “treating” and “treatment” encompasses, e.g., inducing inhibition, regression, or stasis of a disease, disorder or condition, or ameliorating or alleviating a symptom of a disease, disorder or condition. “Ameliorating” or “alleviating” a condition or state as used herein shall mean to relieve or lessen the symptoms of that condition or state. “Inhibition” of disease progression or disease complication in a subject as used herein means preventing or reducing the disease progression and/or disease complication in the subject.
“Administering to the subject” means the giving of, dispensing of, or application of medicines, drugs, or remedies to a subject to relieve, cure, or reduce the symptoms associated with a condition, e.g., a pathological condition.
The drug substance of the present invention, e.g., laquinimod sodium, may be administered in admixture with suitable pharmaceutical diluents, extenders, excipients, or carriers (collectively referred to herein as a pharmaceutically acceptable carrier) suitably selected with respect to the intended form of administration and as consistent with conventional pharmaceutical practices. The unit will be in a form suitable for oral, rectal, topical, intravenous or direct injection or parenteral administration. The compounds can be administered alone or mixed with a pharmaceutically acceptable carrier.
This carrier can be a solid or liquid, and the type of carrier is generally chosen based on the type of administration being used. The active agent can be co-administered in the form of a tablet or capsule, liposome, as an agglomerated powder or in a liquid form. Examples of suitable solid carriers include lactose, sucrose, gelatin and agar. Capsules or tablets can be easily formulated and can be made easy to swallow or chew; other solid forms include granules, and bulk powders.
Capsules or tablets may contain suitable binders, lubricants, disintegrating agents, diluents, coloring agents, flavoring agents, flow-inducing agents, and melting agents. For instance, for oral administration in the dosage unit form of a tablet or capsule, the active drug component can be combined with an oral, non-toxic, pharmaceutically acceptable, inert carrier such as lactose, gelatin, agar, starch, sucrose, glucose, methyl cellulose, magnesium stearate, dicalcium phosphate, calcium sulfate, mannitol, sorbitol and the like.
Suitable binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth, or sodium alginate, povidone, carboxymethylcellulose, polyethylene glycol, waxes, and the like.
Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, stearic acid, sodium stearyl fumarate, talc and the like.
Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum, croscarmellose sodium, sodium starch glycolate and the like.
Specific examples of the techniques, pharmaceutically acceptable carriers and excipients that may be used to formulate oral dosage forms of the present invention are described, e.g., in U.S. Patent Application Publication No. 2005/0192315, PCT International Application Publication Nos. WO 2005/074899, WO 2007/047863, and WO 2007/146248, and in U.S. Pat. No. 7,589,208. For example, the oral dosage form of the present invention may comprise an alkaline-reacting component, said component preferably amounting from about 1 to 20% by weight of the formulation in order to keep the pH above 8.
Techniques and compositions for making dosage forms useful in the present invention are described in the following references: 7 Modern Pharmaceutics, Chapters 9 and 10 (Banker & Rhodes, Editors, 1979); Pharmaceutical Dosage Forms: Tablets (Lieberman et al., 1981); Ansel, Introduction to Pharmaceutical Dosage Forms 2nd Edition (1976); Remington's Pharmaceutical Sciences, 17th ed. (Mack Publishing Company, Easton, Pa., 1985); Advances in Pharmaceutical Sciences (David Ganderton, Trevor Jones, Eds., 1992); Advances in Pharmaceutical Sciences Vol. 7. (David Ganderton, Trevor Jones, James McGinity, Eds., 1995); Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms (Drugs and the Pharmaceutical Sciences, Series 36 (James McGinity, Ed., 1989); Pharmaceutical Particulate Carriers: Therapeutic Applications: Drugs and the Pharmaceutical Sciences, Vol 61 (Alain Rolland, Ed., 1993); Drug Delivery to the Gastrointestinal Tract (Ellis Horwood Books in the Biological Sciences. Series in Pharmaceutical Technology; J. G. Hardy, S. S. Davis, Clive G. Wilson, Eds.); Modem Pharmaceutics Drugs and the Pharmaceutical Sciences, Vol 40 (Gilbert S. Banker, Christopher T. Rhodes, Eds.). All of the aforementioned publications are incorporated by reference herein. Examples of suitable liquid dosage forms include solutions or suspensions in water, pharmaceutically acceptable fats and oils, alcohols or other organic solvents, including esters, emulsions, syrups or elixirs, suspensions, solutions and/or suspensions reconstituted from non-effervescent granules and effervescent preparations reconstituted from effervescent granules. Such liquid dosage forms may contain, for example, suitable solvents, preservatives, emulsifying agents, suspending agents, diluents, sweeteners, thickeners, and melting agents.
Oral dosage forms optionally contain flavorants and coloring agents. Parenteral and intravenous forms may also include minerals and other materials to make them compatible with the type of injection or delivery system chosen.
The compounds used in the method of the present invention may also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamallar vesicles, and multilamellar vesicles. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine, or phosphatidylcholines. The compounds may be administered as components of tissue-targeted emulsions.
The compounds used in the method of the present invention may also be coupled to soluble polymers as targetable drug carriers or as a prodrug. Such polymers include polyvinylpyrrolidone, pyran copolymer, polyhydroxylpropylmethacrylamide-phenol, polyhydroxyethylaspartamidephenol, or polyethyleneoxide-polylysine substituted with palmitoyl residues. Furthermore, the compounds may be coupled to a class of biodegradable polymers useful in achieving controlled release of a drug, for example, polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacylates, and crosslinked or amphipathic block copolymers of hydrogels.
For oral administration in liquid dosage form, the oral drug components are combined with any oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Examples of suitable liquid dosage forms include solutions or suspensions in water, pharmaceutically acceptable fats and oils, alcohols or other organic solvents, including esters, emulsions, syrups or elixirs, suspensions, solutions and/or suspensions reconstituted from non-effervescent granules and effervescent preparations reconstituted from effervescent granules. Such liquid dosage forms may contain, for example, suitable solvents, preservatives, emulsifying agents, suspending agents, diluents, sweeteners, thickeners, and melting agents.
Liquid dosage forms for oral administration can contain coloring and flavoring to increase patient acceptance. In general, water, a suitable oil, saline, aqueous dextrose (glucose), and related sugar solutions and glycols such as propylene glycol or polyethylene glycols are suitable carriers for parenteral solutions. Solutions for parenteral administration preferably contain a water soluble salt of the active ingredient, suitable stabilizing agents, and if necessary, buffer substances. Antioxidizing agents such as sodium bisulfite, sodium sulfite, or ascorbic acid, either alone or combined, are suitable stabilizing agents. Also used are citric acid and its salts and sodium EDTA. In addition, parenteral solutions can contain preservatives, such as benzalkonium chloride, methyl- or propyl-paraben, and chlorobutanol. Suitable pharmaceutical carriers are described in Remington's Pharmaceutical Sciences, Mack Publishing Company, a standard reference text in this field.
The drug substance of the present invention, e.g., laquinimod sodium, may be administered in various forms, including those detailed herein. The treatment with the compound may be a component of a combination therapy or an adjunct therapy, i.e. the subject or patient in need of the drug is treated or given another drug for the disease in conjunction with one or more of the instant compounds. This combination therapy can be sequential therapy where the patient is treated first with one drug and then the other or the two drugs are given simultaneously. These can be administered independently by the same route or by two or more different routes of administration depending on the dosage forms employed.
Gelatin capsules may contain the active ingredient compounds and powdered carriers, such as lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like. Similar diluents can be used to make compressed tablets. Both tablets and capsules can be manufactured as immediate release products or as sustained release products to provide for continuous release of medication over a period of hours. Compressed tablets can be sugar coated or film coated to mask any unpleasant taste and protect the tablet from the atmosphere, or enteric coated for selective disintegration in the gastrointestinal tract.
The compounds used in the method of the present invention may also be administered in intranasal form via use of suitable intranasal vehicles, or via transdermal routes, using those forms of transdermal skin patches well known to those of ordinary skill in that art. To be administered in the form of a transdermal delivery system, the dosage administration will generally be continuous rather than intermittent throughout the dosage regimen.
Parenteral and intravenous forms may also include minerals and other materials to make them compatible with the type of injection or delivery system chosen.
A dosage unit of the compounds used in the method of the present invention may comprise a single compound or mixtures thereof with additional antibacterial agents. The compounds can be administered in oral dosage forms as tablets, capsules, pills, powders, granules, elixirs, tinctures, suspensions, syrups, and emulsions. The compounds may also be administered in intravenous (bolus or infusion), intraperitoneal, subcutaneous, or intramuscular form, or introduced directly, e.g. by injection, topical application, or other methods, into or onto a site of infection, all using dosage forms well known to those of ordinary skill in the pharmaceutical arts.
A “dose” or “dosage unit” of laquinimod as measured in milligrams refers to the milligrams of laquinimod acid present in a preparation, regardless of the form of the preparation. A dosage unit may comprise a single compound or mixtures of compounds thereof. A dosage unit can be prepared for oral dosage forms, such as tablets, capsules, pills, powders, and granules. For example, the “dose” or “dosage unit” of laquinimod may be 0.3, 0.6, or 1.2 mg.
As used herein, a “pharmaceutically acceptable” component is one that is suitable for use with humans and/or animals without undue adverse side effects (such as toxicity, irritation, and allergic response) commensurate with a reasonable benefit/risk ratio.
As used herein, “pharmaceutically acceptable carrier” refers to a carrier or excipient that is suitable for use with humans and/or animals without undue adverse side effects (such as toxicity, irritation, and allergic response) commensurate with a reasonable benefit/risk ratio. It can be a pharmaceutically acceptable solvent, suspending agent or vehicle, for delivering the instant compounds to the subject. The carrier may be liquid or solid and is selected with the planned manner of administration in mind. “Pharmaceutically acceptable carrier” includes “fillers”, which fill out the size of a tablet or capsule, making it practical to produce and convenient for the consumer to use. By increasing the bulk volume, the fillers make it possible for the final product to have the proper volume for patient handling. “Pharmaceutically acceptable carrier” also includes “lubricants”, which prevent ingredients from clumping together and from sticking to the tablet punches or capsule filling machine. Lubricants also ensure that tablet formation and ejection can occur with low friction between the solid and die wall. “Pharmaceutically acceptable carrier” also includes inert carriers such as lactose, gelatin, agar, starch, sucrose, glucose, methyl cellulose, dicalcium phosphate, calcium sulfate, mannitol, sorbitol, microcrystalline cellulose and the like. Liposomes are also a pharmaceutically acceptable carrier.
It is understood that where a parameter range is provided, all integers within that range, and tenths and hundredth thereof, are also provided by the invention. For example, “0.15-0.35%” includes 0.15%, 0.16%, 0.17% etc. up to 0.35%.
The subject invention is also intended to include all isotopes of atoms occurring on the compounds disclosed herein, including impurities. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include tritium and deuterium. Isotopes of carbon including C-13 and C-14.
As used herein, “detection limit” for an analytical method used in screening or testing for the presence of a compound in a sample is a threshold under which the compound in a sample cannot be detected by the analytical method used. The detection limits of a given HPLC method for detecting an impurity in a sample containing laquinimod may vary based on the method and the impurity or impurities being detected. For example, the detection limit of the typical HPLC method for detecting 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid (MCQCA) is 0.03% and the detecting limit of a given method for detecting methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQME), 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline (MCQ), 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (MCQEE) and unknown impurities is 0.02%.
As used herein, “quantitation limit” for an analytical method used in screening or testing for the presence of a compound in a sample is a threshold under which the compound in a sample cannot be quantified by the analytical method used. The quantitation limits of a given HPLC method for detecting an impurity in a sample containing laquinimod may vary based on the impurity or impurities being detected.
As used herein, “density” is a measurement defined as the mass of a substance per unit volume.
As used herein, “bulk density” or “BD” refers to a density measurement of a loose, uncompacted substance, wherein the volume of the substance includes the air trapped between particles.
As used herein, “tapped density” or “TD” refers to a density measurement of a substance that has been tapped or vibrated, thus minimizing the volume of the substance by eliminating or minimizing the air trapped between particles.
As used herein, “rapid stirring” refers to stirring which splashes solvent onto the walls of the vessel.
As used herein, “blend uniformity” refers to the homogeneity of blend or granulate including laquinimod sodium particles prior to encapsulation, tableting or otherwise finalizing the drug product beyond formation of the final blend, and can represent either one sample or the average of more than one sample. Blend uniformity may be measured, for example, by taking 10 samples that represent the upper, middle and lower layer of each batch of the final blend, performing an HPLC assay to measure the amount of active ingredient in the samples, and comparing the amount of active ingredient in each sample to the labeled amount of active ingredient. The standard deviation and relative standard deviation can be determined based on the individual amounts of the tested samples expressed as percentages of the labeled amount of drug substance in each sample.
As used herein, “content uniformity” refers to the homogeneity of the laquinimod sodium content among dosage forms, e.g., capsules or tablets, after formulation. The uniformity of dosage units by content uniformity of the pharmaceutical composition described herein meets the U.S. Pharmacopeia <905> Acceptance Value and range (as specified); L1=15.0 and L2=25.0. Content uniformity may be measured, for example, as indicated by the United States Pharmacopoeia which includes 1) assaying ten tablets (or other dosage form of the drug product) to ensure that the relative standard deviation (RSD) of active content is less than or equal to 6.0% and no value is outside 85-115%; and 2) assaying twenty more tablets (or other dosage form of the drug product) to ensure that the RSD for all thirty is less than or equal to 7.8%, no more than one value is outside 85-115% and no value is outside 75-125% of stated content.
As used herein, “residual solvents” include ethanol, n-heptane, n-octane, methanol, acetone, dioxane, and dimethyl formamide. Residual solvents may be determined, for example, based on the manufacturer's statements of residual solvent levels in the active ingredients/excipients and calculation as per U.S. pharmacopeia <467> Option 2, product meets the USP <467> Residual Solvents limit criteria. Testing is not necessarily required.
As used herein, “NMT” means no more than.
As used herein, “LT” means less than.
As used herein, “MCQME” means methyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate. MCQME is disclosed in U.S. Pat. No. 7,560,557 and J. Org. Chem., 2006, 71, 1658-1667, the contents of which are incorporated by reference into this application. MCQME has the structure:
As used herein, “MCQEE” means ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate. MCQEE is disclosed in U.S. Pat. No. 7,560,557, the contents of which are incorporated by reference into this application. MCQEE has the structure:
As used herein, “MCQ” means 5-chloro-4-hydroxy-1-methylquinolin-2(1H)-one. MCQ is disclosed in U.S. Pat. No. 7,560,557 and J. Org. Chem., 2006, 71, 1658-1667, the contents of which are incorporated by reference into this application. MCQ has the structure:
As used herein, “MCQCA” means 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid. MCQCA is disclosed in U.S. Pat. No. 7,560,557 and J. Org. Chem., 2006, 71, 1658-1667, the contents of which are incorporated by reference into this application. MCQCA has the structure:
As used herein, “NEA” means N-ethyl aniline. NEA is disclosed in U.S. Pat. No. 7,560,557, the contents of which are incorporated by reference into this application. NEA has the structure:
As used herein, “5-HLAQ” means N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. 5-HLAQ is disclosed in PCT International Application No. PCT/US13/26476 and U.S. Application Publication No. US 2013/0217724 A1, the contents of which are incorporated by reference into this application. 5-HLAQ has the structure:
As used herein, “3-HLAQ” means 5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydroquinoline-3-carboxamide. 3-HLAQ is disclosed in PCT International Application No. PCT/US2008/013890 and U.S. Pat. No. 8,178,127 B2, the contents of which are incorporated by reference into this application. 3-HLAQ has the structure:
As used herein, “MEG-LAQ” means N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. MEG-LAQ is disclosed in U.S. Application Publication No. US 2013/0345256 A1, the contents of which are incorporated by reference into this application. MEG-LAQ has the structure:
As used herein, “DELAQ” means 5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. DELAQ is disclosed in PCT International Application No. PCT/US2011/043391 and U.S. Application Publication No. US 2012/0010239 A1, the contents of which are incorporated by reference into this application. DELAQ has the structure:
As used herein, “SPIRO-LAQ” means 1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3′-[l]ethylindolin-[2]-one]. SPIRO-LAQ is disclosed in PCT International Application No. PCT/US2008/013890 and U.S. Pat. No. 8,178,127 B2, the contents of which are incorporated by reference into this application. SPIRO-LAQ has the structure:
As used herein, “BH-3-HLAQ” means 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid. BH-3-HLAQ has the structure:
As used herein, “DMM” means dimethylmalonate. DMM is a synthetic reagent, and has the structure:
As used herein, “DMS” means Dimethyl sulfate. DMS is a synthetic reagent.
As used herein, “MCIA” means 5-chloro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione. MCIA has the structure
As used herein, “DMF” means N,N-dimethylformamide.
As used herein, “LOD” means loss on drying.
The following commercially available solvents of appropriate quality are also used as analytical standards for testing of drug substance: Ethanol, n-heptane, n-octane, methanol, acetone, 1,4-dioxane and N,N-dimethylformamide (DMF). The ID number and source of analytical standards for dimethyl malonate is S4695599 (Merck), sodium methyl sulfate is A0294777 (Arcos), and 13C2-dimethyl sulfate is P-5052 (Holland Moran).
By any range disclosed herein, it is meant that all hundredth, tenth and integer unit amounts within the range are specifically disclosed as part of the invention. Thus, for example, 0.01 mg to 50 mg means that 0.02, 0.03 . . . 0.09; 0.1, 0.2 . . . 0.9; and 1, 2 . . . 49 mg unit amounts are included as embodiments of this invention.
Impurities are measured by common pharmacopeial methods unless otherwise specified.
As used herein, an “anti-solvent” is a solvent in which laquinimod sodium is slightly soluble, very slightly soluble, practically insoluble, or insoluble at room temperature (20-25° C.). The solubility terms are defined below, in accordance with the United States Pharmacopoeia XXV.
The purification of impure crystalline compounds is usually attained by recrystallization from a suitable solvent or mixture of solvents. (Vogel's Textbook of Practical Organic Chemistry. 5′ edition. Longman Scientific & Technical, 1989.) The recrystallization process generally comprises the following steps: a) dissolving the impure crystalline substance in a suitable solvent near the boiling point; b) filtering the hot solution from particles of insoluble material and dust; c) allowing the hot solution to cool to cause the dissolved substance to crystallize out; and d) separating the crystals from the supernatant solution. (Id.) However, standard recrystallization techniques were accompanied by low or no yields when applied to laquinimod sodium as taught in U.S. Pat. No. 7,884,208. As shown in Examples 2-7 of U.S. Pat. No. 7,884,208, attempts to recrystallize laquinimod sodium using standard recrystallization procedures resulted in poor yields, if any. The process of U.S. Pat. No. 7,884,208 overcomes the difficulties associated with recrystallizing laquinimod sodium by use of an anti-solvent in which laquinimod sodium is practically insoluble. In addition, the process of U.S. Pat. No. 7,884,208 concentrates the laquinimod sodium aqueous solution before the addition of the anti-solvent. The process of the present invention is an improvement over the process of U.S. Pat. No. 7,884,208.
This invention will be better understood by reference to the Experimental Details and Examples which follow, but those skilled in the art will readily appreciate that the specific experiments detailed are only illustrative of the invention as described more fully in the claims which follow thereafter.
The particle size distributions were measured by Malvern Laser Diffraction, using the Mastersizer S model. Laser diffraction relies on the fact that diffraction angle of light is inversely proportional to particle size. Properties of particles are measured and interpreted as measurements of a sphere (a sphere being the only shape that can be described by one unique number). In addition, laser diffraction calculates a particle size distribution based around volume terms, thus eliminating particle count from the determination of particle size. The Mastersizer S model measures particles using a single technique and a single range setting.
D(0.1) is the particle size, in microns, below which 10% by volume distribution of the population is found. D(0.5) is the particle size, in microns, below which 50% by volume distribution of the population is found. D(0.9) is the particle size, in microns, below which 90% by volume distribution of the population is found.
Metal content was measured using inductively coupled plasma atomic emission spectrometry using an inductively coupled plasma atomic emission spectrometry (“ICP-AES”) system manufactured by Spectro (Kleve, Germany). Sample digestion was performed in 65% nitric acid, and the internal standard used was scandium.
Note: In the following examples the volumes of solvents used are calculated relative to starting weight of laquinimod sodium. The yields are calculated in weight percent.
Laquinimod sodium and polar impurity/degradation products were determined by isocratic reversed phase high performance liquid chromatography (RP-HPLC), using an ODS-3V column and a mobile phase comprised of a mixture of ammonium acetate buffer at pH 7.0 (80%) and acetonitrile (20%). The detection technique was ultraviolet absorption at 240 nm.
Weigh accurately in duplicate about 15 mg of laquinimod sodium standard into a 50 mL volumetric flask. Dilute with diluents A up to ⅔ of the volume, sonicate for 2 minutes in a cold sonication bath and dilute to volume with diluents A.
Concentration of standard stock solution is about 300 μg/mL laquinimod sodium. Standard stock solution may be used for one month when stored in a refrigerator 2° C.-8° C.
Dilute 3 mL of the Standard Stock Solution to 10 mL with diluents B (dilution factor 3.33).
Concentration of Laquinimod sodium is about 90 μg/mL. Concentration expressed as laquinimod (acid) is about 85 μg/mL.
Standard working solution A may be used for 7 days when stored in a refrigerator (2° C.-8° C.).
Weigh accurately about 18 mg of MCQCA standard into a 100 mL volumetric flask. Dilute to volume with acetonitrile, sonicate (in a cold sonication bath) until the substance is completely dissolved—stock MCQCA solution.
Concentration of MCQCA is about 180 μg/mL.
MCQCA Stock standard solution should be freshly prepared.
Prepare a solution in diluents B, containing Laquinimod in a concentration of 0.2% and MCQCA—in a concentration of 0.1%, with respect to the working concentration of Laquinimod in Standard solution A. As an example, apply the following procedure.
Transfer 4.0 mL of laquinimod sodium standard solution for assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a 100 mL volumetric flask and dilute to volume with the diluents B (intermediate dilution).
Place 2.5 mL of this intermediate dilution into a 50 mL volumetric flask and make up to volume with diluents B.
Total dilution factor for laquinimod standard is 1666.67, for MCQCA 2000.
Concentration of laquinimod sodium is about 0.18 μg/mL (0.2%)
Concentration of MCQCA is about 0.09 μg/mL (0.1%, QL level).
Standard solution I may be used for 24 hours when stored in a refrigerator.
Prepare solution containing the following potential impurities standards (markers) using the Diluent A as a solvent:
MCQ: 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
MCQCA: 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid
MCQMA: Methyl 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
5-HLAQ: N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide
The Mixed Solution may be prepared as follows:
Weigh about 3 mg of each impurity standard/marker into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A.
Concentration of each impurity in the Mixed Solution is about 30 μg/mL. Mixed Solution may be used for up to 4 months when stored frozen at about −20° C. For this purpose, the freshly prepared Mixed Solution should be divided into aliquots, immediately frozen and stored at −20° C. After thawing, the aliquots should be mixed well and should not be refrozen.
Weigh about 3 mg of MCQEE (Ethyl 5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate) into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A. This is MCQEE Stock solution.
This solution may be used for up to 4 months when stored frozen at about −20° C.
For this purpose, the freshly prepared MCQEE Stock Solution should be divided into aliquot, immediately frozen and stored at −20° C. After thawing, the aliquots should be mixed well and should not be refrozen.
Weigh about 3 mg of MEG-LAQ (Meglumine Adduct of Laquinimod) into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A. This is MEG-LAQ Stock solution.
This solution may be used for one week when stored in refrigerator (2° C.-8° C.).
Prepare two Resolution Solutions separately as follows, using the Mixed Solution.
Transfer 3 mL of laquinimod standard stock solution (solution S), 0.3 mL of the Mixed Solution and 0.3 mL of the MCQEE Stock solution to a 10 mL volumetric flask and dilute to volume with the Diluent B. This is Resolution Solution 1.
Concentration of laquinimod sodium in it is about 90 μg/mL. Concentration of each impurity—is about 0.9 μg/mL (about 1% with respect to the working concentration of laquinimod).
Resolution Solution 1 is used for resolution test (for system suitability) and for determination of retention times (RT)/relative retention times (RRT) of five impurities: MCQ, MCQCA, MCQME, MCQEE, and 5-HLAQ.
Resolution Solution 1 may be used for 9 days if stored in a refrigerator 2° C.-8° C.).
Transfer 3 mL of laquinimod standard stock solution (solution S), 0.3 mL of the Mixed Solution and 0.3 mL of the MEG-LAQ Stock solution to a 10 mL volumetric flask and dilute to volume with the Diluent B. This is Resolution Solution 2.
Concentration of laquinimod sodium in it is about 90 μg/mL. Concentration of each impurity—is about 0.9 μg/mL (about 1% with respect to the working concentration of laquinimod).
Resolution Solution 2 is used for determination of retention time of MEG-LAQ.
Resolution Solution 2 may be used for 9 days if stored in a refrigerator 2° C.-8° C.).
Weigh accurately 20 capsules and completely empty their contents into a mortar. Pay attention to complete emptying of capsule contents into the mortar, using spatula when necessary. Weigh the empty capsules. Calculate the average weight of the capsule contents. Mix and grind the capsule contents in a mortar and keep the powder in a tightly closed container protected from light.
Weigh accurately, in duplicate, the amount of powder corresponding to 7 capsules, into a 50 mL volumetric flask.
Add diluents B up to ⅔ of the volume, shake for 30 minutes at 200 mot/min. Dilute to volume with Diluent B. Mix well. Filter before use through a 0.45 μm GHP ACRODISC GF filter or equivalent, discarding the first 0.5-1 mL.
Working concentration of laquinimod (acid) is about 84 ug/mL. Immediately after preparation place sample solutions into a refrigerator or in a cooled to 5° C. autosampler. The sample solutions may be used for 24 hours when kept at the temperature 2° C.-8° C.
Inject the Resolution Solutions, Diluent B (Blank), Standard Solutions for assay and IDD and Sample solutions, according to standard operating procedures.
Determine the retention time (RT) and the area of the laquinimod peak in the chromatograms of Sample and Standard Solutions for identification and assay.
Determine the RT, the relative retention time (RRT) and peak areas of all known impurities and any other impurities in the chromatograms of Sample Solutions, for calculation of the content of impurities/degradation products.
Ignore early elution peaks of excipients and system peaks (See chromatogram for determination of impurities/degradation products). For example. Use integration inhibition between 0 and RRT 0.15 (about 2.5 minutes).
Adjust integration parameters to reject peaks with area less than 10% of the average response of Laquinimod peak in the injections of Standard Solution I (for determination of impurities/degradation products).
Disregard peak of MEG-LAQ in sample injections (identified following Resolution Solution 2). The content of MEG-LAQ is tested by Example 23.
Typical retention time of laquinimod peak is 15.5±2.0 minutes.
Tailing factor (USP) for laquinimod peak should be not more than 2.0.
Resolution factor for all the pairs of peaks should be not less than (NLT) 2.
RRT of the peaks of known impurities/degradation products should be as follows:
MCQME: 0.33-0.38
MCQ: 0.49-0.58
MCQEE: 0.56-0.65
MCQCA: 0.71-0.85
5-HLAQ: 1.2-1.4 (Should not be more than 23 minutes)
MEG-LAQ peak is substantially broadened in comparison with neighboring peaks. Retention time of MEG-LAQ is variable, being very sensitive to slightest changes in chromatographic conditions (pH, % acetonitrile, Temperature, etc.) and therefore should be defined using its peak in the chromatogram of the Resolution Solution 2. Typically, its RRT is about 0.66.
Evaluate laquinimod standards for assay and IDD in order to test the system precision according to the standard operating procedures.
MCQCA in Solution I is used to test the sensitivity of the system. RSD of the area of six injections of Std 1 as well as the difference between Std 1 and Std 2 should be no more than (NMT) 20%.
Injection diluents B to detect system peaks.
The RT of the main peak obtained in the sample chromatogram should correspond to that obtained for the laquinimod peak in the injection of Standard Solution.
Where 0.94 is the conversion factor of laquinimod sodium salt to laquinimod (acid).
Areaimpurity is the area of an impurity/degradation product (known or unknown) peak in the Sample Solution.
Areastd is the laquinimod peak in chromatogram of Standard Solution I.
0.94 is the conversion factor of laquinimod sodium salt to laquinimod (acid).
RRF is the relative response factors of impurities/degradation products calculated as the following ratio: slope of Laquinimod regression line/slope of impurity regression line.
The values for relative response factors with respect to laquinimod are: MCQME: 0.74; MCQ: 0.65; MCQEE: 0.85; MCQCA: 0.62; and 5-HLAQ: 1.0.
RRF for unknown impurities/degradation products is taken as 1.0.
7.2.3 Evaluation and Report of Impurities/Degradation products
Quantitation level (QL) MCQME, MCQ, MCQEE, 5-HLAQ, and unknown impurities is 005%. Detection level (DL) of MCQME, MCQ, MCQEE, 5-HLAQ and unknown impurities is 0.02%. QL of MCQCA is 0.1%. Detection level DL of MCQCA is 0.03%.
Correlate all the peaks in sample chromatogram with those in the system suitability chromatogram, with ±5% of the actual corresponding retention times. Report data as shown in Table 2.
Recrystallization of laquinimod sodium was performed on pilot scale (Batches A and B) as follows.
Re-crystallization of Laquinimod Na on pilot scale is performed in two glass-lined reactors (Reactor A, 30 liter volume and Reactor B 60 liter). Solid product is filtered and dried in Hastelloy C agitated filter-dryer with 20 micron mesh.
Batch size is 2.5 kg of starting crude Laquinimod Na.
Batch of crude Laquinimod Na (2.5 kg) is introduced to Reactor A with 10 volumes of process water. The batch is heated to 60-73° C. at stirring until complete dissolution of solid.
The hot solution in Reactor A is transferred to Reactor B through 0.2 μm filtration system. Reactor A and filters washed with 1.2 volumes of process water and the wash is transferred to the Reactor B.
Vacuum is built-up and the solution in the Reactor B is evaporated at P<45 mmHg and jacket temperature T<65° C. until volume of the residue reaches 5.4 liters (2.16 volumes). Then atmospheric pressure is built-up and jacket temperature 40-50° C. is adjusted. The batch is stirred for not less than 10 minutes and then seeded with Laquinimod Na crystals to initiate crystallization.
The batch is stirred at 45° C. for additional 90 minutes and 7.9 volumes of acetone are added to the reactor in 1.5-2.5 hrs. Reactor temperature during the addition maintained between 40 and 50° C.
Resulting slurry is cooled to 0±4° C. during 3.5-4.5 hrs and stirred at this temperature for 10-15 hrs. Then the slurry is transferred to filter-dryer and solid is filtered under pressure of nitrogen.
The cake is washed twice (2·2 kg) with acetone, purged with nitrogen and then dried under vacuum (P<50 mmHg) and elevated temperature (T=40° C.) at agitation.
Dry product is discharged, sampled for analysis and packed.
The pilot scale process of recrystallization of laquinimod sodium was based on Example 15 of U.S. Pat. No. 7,884,208. The starting material was crude laquinimod sodium having low particle size (d(0.1)=1-2μ, d(0.5)=6-11μ; d(0.9)=20-35μ) and appears as aggregated solid. Example 15 of U.S. Pat. No. 7,884,208 involves 25.0 g of laquinimod sodium (laboratory scale) prepared according to the method disclosed in U.S. Pat. No. 6,875,869. In Example 15, the 25.0 g of laquinimod sodium is dissolved in an aqueous solution of laquinimod sodium and then evaporated under vacuum at stirring to a concentrated solution having a volume ratio of 2.14 v/w, the resulting residue is seeded to induce crystallization then treated with an anti-solvent (acetone).
The modified pilot scale process was performed with 2.5 kg of laquinimod sodium which is a 100-fold scale up from Example 15. In addition, the modified pilot scale process had significant differences from the laboratory scale process of Example 15 of U.S. Pat. No. 7,884,208. Specifically, evaporation on the laboratory scale was performed in a round-bottom flask in a rotary evaporator without stirring, while evaporation on the pilot scale was performed in a reactor with stirring. On the pilot scale, the evaporation residue is stirred aggressively, liquid splashes on the reactor walls, solid depositions form, and crystallization was spontaneous. On the laboratory scale, a metastable solution could be concentrated to a volume ratio of 2.1-2.2 v/w at which point crystallization did not take place and nucleation was controlled by seeding. On the pilot scale, conditions and concentration were such that spontaneous crystallization took place, i.e., crystallization was induced without seeding.
Surprisingly, the pilot batches did not result in laquinimod sodium particles having a particle size distribution expected based on Example 15 of U.S. Pat. No. 7,884,208. Instead, applicants unexpected found that the pilot batches resulted in a mixture of recrystallized laquinimod sodium particles wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 40 microns, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 15 microns, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 5 microns. A comparison of particle size distributions resulting from the two different processes is shown in Table 4.
The process of U.S. Pat. No. 7,884,208 and the process of Example 1, above, each produce different products and are not equivalent processes. Applicants' pilot scale version of the process of U.S. Pat. No. 7,884,208 resulted in substantially different conditions from the prior art and resulted in a substantially different product having smaller particle sizes as shown in Table 4.
Since laquinimod Na is a potent drug substance, small particle size is advantageous for this API. Formation of non-aggregated laquinimod sodium crystals with reduced particle size could provide better uniformity of drug product and avoid milling or de-lumping operations. The starting material, crude laquinimod sodium, appears as aggregated solid. The re-crystallized product is free flowing powder. Powders with smaller particles have a stronger trend to aggregate. Crude laquinimod sodium prepared by slurry-to-slurry recrystallization (i.e., the process of U.S. Pat. No. 6,077,851) produces particles having a low particle size and are highly aggregated. The modified process produces particles having a low particle size and are free flowing.
The aim was scalable crystallization procedure giving smaller crystal size, PSD and low aggregation on laboratory, pilot and commercial scale. The desirable PSD profile was the following: d(0.1)<5 μm, d(0.5)<15 μm and d(0.9)<40 μm. The method is based on spontaneous crystallization initiated in aqueous phase prior to acetone addition. The important factor affecting crystallization is initial concentration of crystallizing solution. In the new crystallization procedure, reduced water volume ratio in the end of evaporation from 2.14 v/w to 1.7-1.8 v/w. Higher concentration of the solution ensures initiation of spontaneous crystallization in the end of evaporation operation and provides higher supersaturation level and lower crystal size.
Recrystallization of laquinimod sodium was performed on laboratory scale (Batch A) as follows.
All operations of Laquinimod Na re-crystallization step including evaporation were performed on laboratory scale in transparent agitated glass reactors equipped with stirrer, thermometers and circulating bath for heating and cooling.
25 g crude Na Laquinimod and 275 ml deionized water introduced into 250 ml stirred jacketed glass reactor. The mixture is stirred and heated to 70° C., after complete dissolution of the solid the solution is filtered through paper filter. Resulting clear filtrate introduced to 250 ml jacketed glass reactor equipped with circulating bath, stirrer, thermometer and vacuum distillation system.
Vacuum is applied and water is distilled at stirring, pressure during the evaporation is 38-40 mbar and jacket temperature is 55° C.
After distillation of ca. ⅔ volume spontaneous crystallization on the reactor wall above liquid level is observed.
The distillation is continued until the residue volume reaches 45 ml then atmospheric pressure is build up and the batch is stirred at 50° C. for one hour. On this step intensive crystallization takes place.
200 ml acetone is added to the resulting slurry in one hour and the batch is stirred for one additional hour at 50° C.
The batch is cooled to 0-5° C. during one hour and filtered on Büchner filter. The solid cake is washed with 75 ml of acetone.
Collected wet product (28.0 g) is dried in oven under vacuum at 50° C. to constant weight.
Dry product—23.8 g; Crystallization yield—95.2%
Microscopic observation—rod-shape particles
Particle Size Distribution by Malvern:
D(0.1)=2.3 μm; D(0.5)=10.8 μm; D(0.9)=32.7 μm
Recrystallization of laquinimod sodium was performed on laboratory scale (Batch B) as follows.
All operations of Laquinimod Na re-crystallization step including evaporation were performed on laboratory scale in transparent agitated glass reactors equipped with stirrer, thermometers and circulating bath for heating and cooling.
25 g Na Laquinimod crude and 275 ml deionized water introduced into 250 ml stirred jacketed glass reactor. The mixture is stirred and heated to 70° C., after complete dissolution of the solid the solution is filtered through paper filter. Resulting clear filtrate introduced to 250 ml jacketed glass reactor equipped with circulating bath, stirrer, thermometer and vacuum distillation system. Vacuum is applied and water is distilled at stirring, pressure during the evaporation is 38-40 mbar and jacket temperature is 55° C. During the distillation spontaneous crystallization on the reactor wall is observed when the residue volume reached ca. 120 ml. The distillation is continued until the residue volume reaches 45 ml then atmospheric pressure is build up and the batch is stirred at 50° C. for one hour. On this step intensive crystallization takes place.
200 ml acetone is added to the resulting slurry in one hour and the batch is stirred for one additional hour at 50° C.
The batch is cooled to 0-5° C. during one hour, stirred at this temperature for one additional hour and filtered on Büchner filter. The solid cake is washed with 75 ml of acetone.
Collected wet product (27.5 g) is dried in oven under vacuum at 50° C. to constant weight.
Dry product—23.65 g; Crystallization yield—94.6%
Microscopic observation—rod-shape particles
Particle Size Distribution by Malvern:
D(0.1)=2.6 μm; D(0.5)=12.4 μm; D(0.9)=34.3 μm
25 g
The results of Example 2 and Example 3 are summarized in Table 5. Table 5 shows that the process reliably produced a mixture of laquinimod sodium crystals a mixture of recrystallized laquinimod sodium particles wherein (i) 90% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 40 microns, (ii) 50% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 15 microns, and (iii) 10% or more of the total amount by volume of the laquinimod sodium particles have a size of less than 5 microns.
Recrystallization of laquinimod sodium was performed on a production scale (Batches C, D and E) as follows.
Re-crystallization of Laquinimod Na on the production scale is performed in two 250 liter glass-lined reactors (Reactor I and Reactor II). Solid product is filtered and dried in Hastelloy C-22 agitated filter-dryer with 20 micron mesh.
Batch size is 6.5-7.5 kg of dry API.
Batch of crude Laquinimod Na is introduced to Reactor I with 11 volumes of process water. The batch is heated to 60-73° C. at stirring until complete dissolution of solid.
The hot solution in Reactor I is circulated through 0.2 μm filtration system at heating and stirring during 15-20 minutes. After the circulation completion filtered solution is transferred to Reactor II through 0.2 μm filter. Reactor I and filters washed with 1.75 volumes of process water and the wash is transferred to the Reactor II.
Vacuum is build-up and the solution in the Reactor II is evaporated at P<45 mmHg and jacket temperature T<65° C. until volume of the residue reaches 14-16 liter (ca. 1.7-1.8 v/w water/weight crude laquinimod Na starting material). On this step spontaneous crystallization is initiated on the reactor walls. Then atmospheric pressure is build-up and jacket temperature 40-50° C. is adjusted.
The batch is stirred for not less than 10 minutes.
The batch is stirred at 45° C. for additional 90 minutes and 7.9 volumes of acetone are added to the reactor in 1.5-2.5 hrs. Reactor temperature during the addition maintained between 40 and 50° C.
Resulting slurry is cooled to 0±4° C. during 2-5 hrs and stirred at this temperature for 10-15 hrs. Then the slurry is transferred to filter-dryer and solid is filtered under pressure of nitrogen.
The cake is washed twice (2×10 liter) with acetone, purged with nitrogen and then dried under vacuum (P<50 mmHg) and elevated temperature (T=35±5° C.) at agitation.
Dry product is discharged, sampled for analysis and packed.
Data for 3 typical production GMP batches is summarized in Table 6. The PSD values presented in Table 6 are in a good accordance to the results of laboratory scale experiments presented in Example 2 and Example 3.
Batch C and D had reduced levels of impurities (Table 7) and good bulk and tapped density (Table 6).
Analysis of powder density of laquinimod sodium produced on the production scale (22 crystallization batches) shows that the bulk density varies in range between 0.237 and 0.364 g/ml. Tapped density is with 0.432 and 0.609 g/ml.
The results of analyzing Batches C and D are shown in Table 7 and Table 8.
Escherica Coli
Microscopic photographs of typical batches of Crude and re-crystallized Laquinimod Na at different magnification are presented on
The modified crystallization procedure demonstrated good reproducibility of particle size distribution on the production scale. Reduction of the evaporation residue volume to a ratio of 1.7-1.8 v/w and initiation of spontaneous crystallization provided desirable crystal size. Improved crystallization procedure demonstrates reduction in crystal size to a level of d(0.9)<40 μm and good reproducibility of Particle Size Distribution on the production scale.
The product with reduced crystal size has no trend to aggregation and does not need milling or de-lumping for homogenization. The product with similar PSD prepared by slurry-to-slurry procedure is aggregated and thus problematic in formulation.
The modified crystallization procedure also results in laquinimod sodium having desirable density and purity profiles.
The improved crystallization procedure also provides effective purification from organic impurities, e.g., MCQME.
The data shown in Table 9 demonstrates complete removal of MCQME impurity by re-crystallization of commercial scale batch of Laquinimod sodium. Since this intermediate has genotoxic potential it should be purified to undetectable level. The crystallization process also provides purification of all other known organic impurities to the level below the limit of detection.
Microscopic photographs of typical batches of crude and re-crystallized Laquinimod sodium at different magnification are presented on
Laquinimod capsules are manufactured according to the method as described in Example 2 of PCT International Application Publication No. WO 2007/146248, the entire content of which is hereby incorporated by reference. Steps of Example 2 of WO 2007/146248 are performed. Each capsule contains 064 mg of laquinimod sodium equivalent to 0.6 mg laquinimod.
The capsules had quantities of impurities within specification based on HPLC relative to the amount of laquinimod.
The capsules have a water content of no more than 1.5%.
The dissolution profiles, content uniformity, and residual solvents of the encapsulated pharmaceutical composition conforms to U.S. Pharmacopeia <711> (dissolution), U.S. Pharmacopeia <905> (uniformity), and U.S. Pharmacopeia <467>.
Each capsule contains 90.0-110.0% of the labeled amount.
The capsules contain a total aerobic microbial count (TAMC) of NMT 103 cfulg, a total combined yeasts/moulds count (TYMC) of NMT 102 cfu/g, and an absence of Escherichia Coli in 1 g.
Example 5 demonstrates that, in a commercial-scale production, pharmaceutical compositions of laquinimod can be prepared with non-detectable levels or a low level of polar impurities and non-polar impurities.
Laquinimod sodium primary reference standard batch was prepared by crystallization of laquinimod sodium batch, followed by purification by crystallization from a water/acetone mixture (1:4.5 w/w). The crystallization was performed by dissolving the drug substance batch in water while heating. The clear solution was filtered and concentrated to a known residue volume under reduced pressure. Acetone was added and the solution was cooled. The obtained crystals were filtered, washed and dried. The chromatographic purity of this batch was found to be 100.0%. The total of both polar and non-polar impurities was LT 0.05%, content of DELAQ was LT 0.1%, and laquinimod acid content was LT 0.2%.
MCQ is a white solid. The molecular structure, chemical formula and molecular weight of MCQ are provided below.
MCQ was prepared in the following manner: MCQCA and DMSO were heated at 75° C. for 2 hours and were then cooled to room temperature. Water was added and the precipitate was collected by filtration, washed with water and dried in a vacuum oven at 50° C. Material of sufficient purity to be suitable for use as a reference standard was obtained.
MCQ was characterized by NMR, MS, elemental analysis and FT-IR as detailed hereafter.
The 1H-NMR and 13C-NMR characterization of MCQ was performed in DMSO on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 10. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for MCQ
13C (ppm)
1H (ppm)
The mass spectrum of MCQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. The results for MCQ are presented in Table 12. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%.
The FT-IR spectrum of MCQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
MCQCA is a white solid. The molecular structure, chemical formula and molecular weight of MCQCA are provided below.
MCQCA was prepared in the following manner:
A mixture of MCQEE in a 2.8M solution of HCl in acetic acid was heated for 6 hours at 65° C. using a reflux condenser. The mixture was cooled to room temperature, diluted with 2-propanol and was further cooled to 8° C. The crystals were filtered off, washed with 2-propanol and dried in a vacuum oven at 50° C.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
MCQCA was characterized by NMR, MS, elemental analysis and FT-IR as detailed below.
The 1H-NMR and 13C-NMR characterization of MCQCA was performed in D2O+KOH on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 14. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for MCQCA
13C (ppm)
1H (ppm)
The mass spectrum of MCQCA was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%.
The results for MCQCA are presented in Table 16.
The attenuated total reflectance (ATR) FT-IR spectrum of MCQCA was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
MCQME is a white to yellowish solid. The molecular structure, chemical formula and molecular weight of MCQME are provided below.
The preparation of MCQME started with the formation of sodium dimethylmalonate by reaction of dimethylmalonate in DMF with sodium methoxide solution. Then, the intermediate MCIA was reacted with sodium dimethylmalonate to form the MCQME sodium salt. Finally the MCQME sodium salt was acidified to MCQME, which was washed with water and dried. The following is a detailed description of the preparation of MCQME.
DMF (66.5 kg [70.4 liter]), dimethyl malonate (9.6 kg [72.7 mole, 8.3 liter]) and a 30% methanolic solution of sodium methoxide (1.5 kg [63.9 mole]) were charged to a reactor. The reaction mixture was stirred and heated to 82° C. to 88° C. and kept at this temperature for 1½ to 2½ hours in order to form sodium dimethyl malonate. Then the mixture was cooled to 30-40° C. Vacuum was applied and the reactor jacket temperature was heated gradually in order to evaporate out methanol. After evaporation of methanol (not less than 12 kg [14.5 liter] distillate) the vacuum was broken with nitrogen and the reaction mixture was heated to 82° C. to 88° C. C. Then MCIA (8.5 kg [40.1 mole]) was added gradually to the reaction mixture and washed with DMF (10 kg [10.6 liter]). The reactor content was cooled to less than 75° C., vacuum was applied, the reactor jacket temperature was gradually raised and methanol was evaporated (12 kg [15 liter] distillate). The vacuum was broken with nitrogen and the reaction mixture was cooled to 12° C. to 35° C. Process water (119 Kg) was added gradually, keeping the reaction mixture temperature at 12° C. to 35° C., then the reaction mixture was heated to 42-48° C. 32% Aqueous HCl solution (11.1 kg [97.3 mole]) was added during 2 to 3 hours and remains of hydrochloric solution in feeding line were washed out with DMF. The formed slurry is cooled to 20° C. to 30° C. and was maintained at that temperature for not less than 30 minutes.
The slurry was transferred gradually to a filter-dryer. Mother liquor was removed by pressing the slurry. Process water (29.8 kg for each cycle) was added to the filter-dryer and pressed out. Washing of the filter-dryer content with water was performed until the pH of outflow washing liquor was NLT 4. The product was dried under vacuum at 40° C. to 50° C. (jacket temperature of filterdryer).
Vacuum drying was performed until the LOD of a sample from the filter-dryer was less than 2.0% and the water content (by Karl-Fischer) was not more than 0.5%. Then the jacket was cooled to 20° C. to 30° C. and dry MCQME was discharged.
The yield is 62% to 87%.
The dry MCQME was delumped, analyzed and released.
MCQEE is a light yellow solid. The molecular structure, chemical formula and molecular weight of MCQEE are provided below.
MCQEE was prepared in the following manner:
Sodium hydride was added to a solution of MCIA and diethyl malonate in DMF. The mixture was gradually heated to 95° C. and stirred for 3.5 hours, then cooled to 35° C. The reaction mixture was quenched with water and then acidified with 37% HCl. After 1 hour at 8° C. the mixture was filtered and washed with water (until the pH of the mother liquor was 4.5) and dried in a vacuum oven at 40° C. The crude product was crystallized by dissolving in ethanol at 85° C. followed by slow cooling (in an ice water bath). The obtained crystals were filtered, washed with ethanol and dried in vacuum oven at 50° C.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
MCQEE was characterized by NMR, MS, elemental analysis and FT-IR as detailed below.
The 1H-NMR and 13C-NMR characterization of MCQEE was performed in CDCl3 on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 18. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for MCQEE
13C (ppm)
1H (ppm)
The mass spectrum of MCQEE was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for MCQEE are presented in Table 20.
The attenuated total reflectance (ATR) FT-IR spectrum of MCQEE was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus, A representative spectrum is presented in
5-HLAQ is an off-white solid. The molecular structure, chemical formula and molecular weight of 5-HLAQ are provided below.
5-HLAQ is unstable in most organic solvents as well as in aqueous acidic medium. Therefore purification by the commonly used methods is extremely difficult. Consequently, purification of the material was performed by formation of the di-acetate derivative followed by basic hydrolysis, acidic precipitation and rapid filtration.
5-HLAQ was prepared by a multi-step procedure, depicted below.
The synthesis involved: 1) preparation of 5-MeO-laquinimod (compound 2) from 2-amino-6-methoxybenzoic acid (compound 1); 2) preparation of 5-HLAQ crude (compound 3); 3) acetylation of the phenolic groups to produce the di-acetate-laquinimod derivative (compound 4); 4) hydrolysis of the di-acetate derivative to produce the purified 5-HLAQ (compound 5).
The procedure for the preparation of 5-HLAQ is further described step-wise in the following paragraphs:
5-MeO-laquinimod was obtained by a four-step synthetic procedure starting from 2-amino-6-methoxybenzoic acid (compound 1). These four steps are identical to the manufacturing process of laquinimod starting from ACBA.
5-MeO-laquinimod was dissolved in dichloromethane and the solution was cooled to 0-5° C. AlCl3 was added and the reaction mixture was stirred for 0.5 hour at 0-5° C. followed by 7 hours' stirring at ambient temperature. The solution was evaporated to dryness at 30° C. (in a water bath) and water was added. The obtained grey solid was filtered, washed with 1N HCl and dried at 30° C. in a vacuum oven.
Acetic anhydride was added to a solution of 5-HLAQ crude in pyridine and the reaction mixture was stirred for 1 hour at room temperature. The pyridine was evaporated to dryness and the oily residue was dissolved in dichloromethane. The organic solution was washed with 1N HCl followed by aqueous washings. The crude 2 was purified by flash chromatography on silica gel (mobile phase: 1% methanol in dichloromethane).
1N NaOH solution was added to a suspension of 4 in ethanol. The reaction mixture was stirred for 40 min and the ethanol was evaporated. The residue was acidified with 5N HCl down to pH 1-2 and the white solid was filtered, washed with water and dried. 5-HLAQ was obtained as a white solid with a purity of >99% by HPLC.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
5-HLAQ was characterized by NMR, MS, elemental analysis and FT-IR as detailed below.
The 1H-NMR and 13C-NMR characterization of 5-HLAQ was performed in DMSO on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 22. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for 5-HLAQ
13C (ppm)
1H (ppm)
The mass spectrum of 5-HLAQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+).
The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for 5-HLAQ are presented in Table 24.
The attenuated total reflectance (ATR) FT-IR spectrum of 5-HLAQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
DELAQ is an off-white to pale yellow solid. The molecular structure, chemical formula and molecular weight of DELAQ are provided below.
DELAQ was prepared in the following manner:
A mixture of MCQME, n-heptane, n-octane and aniline was heated in a jacketed reactor to Tj=135° C. and the volatiles were distilled for 3 hours. The distillation was stopped and additional portions of n-heptane and aniline were added to the mixture. The distillation was continued for an additional 2 hours at Tj=135° C. The reaction mixture was cooled to room temperature and n-heptane was added. The crystalline solid formed was collected by filtration and washed with n-heptane. The wet material was dried in a vacuum oven at 50° C.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
DELAQ was characterized by NMR, MS, elemental analysis and FT-IR, as detailed below.
The 1H-NMR and 13C-NMR characterization of DELAQ was performed in CDCl3 on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 26. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for DELAQ
13C (ppm)
1H (ppm)
The mass spectrum of DELAQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for DELAQ are presented in Table 28.
The FT-IR spectrum of DELAQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
3-HLAQ is a yellow solid. The molecular structure, chemical formula and molecular weight of 3-HLAQ are provided below.
3-HLAQ was prepared in the following manner:
A solution of laquinimod sodium in water was added drop-wise to a solution of disodium hydrogenphosphate dihydrate and oxone (2KHSO5.KHSO4.K2SO4) in water. 10% NaOH was added until the pH was 8.0. The solution was mixed for 30 minutes at room temperature. The obtained solid was filtered, washed with water and dried in a vacuum oven at 50° C. The crude product was recrystallized twice from ethyl acetate and n-heptane mixture. Each time the product was isolated by cooling the mixture to 10° C. and stirring for 1 hour. The obtained crystals were filtered off, washed with n-heptane and dried in a vacuum oven at 50° C.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
3-HLAQ was characterized by NMR, MS, elemental analysis and FT-IR, as detailed below.
The 1H-NMR and 13C-NMR characterization of 3-HLAQ was performed in CDCl3 on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 30. The 1H-NMR and 13C-NMR spectra are presented in
1H and 13C -NMR Peak Assignments for 3-HLAQ
13C (ppm)
1H (ppm)
The mass spectrum of 3-HLAQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for 3-HLAQ are presented in Table 32.
The attenuated total reflectance (ATR) FT-IR spectrum of 3-HLAQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
SPIRO-LAQ is a white powder. The molecular structure, chemical formula and molecular weight of SPIRO-LAQ are provided below.
SPIRO-LAQ was prepared in the following manner:
A mixture of laquinimod (free acid), ammonium cerium (IV) nitrate, ethanol and acetic acid in water was mixed for 1 hour. Then the precipitate was washed and filtered with water followed by ethanol, and dried in a vacuum oven at 50° C.
Material of sufficient purity to be suitable for use as a reference standard was obtained.
SPIRO-LAQ was characterized by NMR, MS, elemental analysis and FT-IR as detailed below.
The 1H-NMR and 13C-NMR characterization of SPIRO-LAQ was performed in DMSO on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 34. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for SPIRO-LAQ
13C (ppm)
1H (ppm)
The mass spectrum of SPIRO-LAQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The ratio between MS signals containing 35Cl and 37Cl corresponds to the natural abundances of these isotopes (about 3:1). The spectrum shown in
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for SPIRO-LAQ are presented in Table 36.
The attenuated total reflectance (ATR) FT-IR spectrum of SPIRO-LAQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus. A representative spectrum is presented in
The molecular structure, chemical formula and molecular weight of MEG-LAQ is presented below.
An aqueous solution of laquinimod sodium and meglumine was refluxed and bubbled with air for about 1 month. The obtained solution was diluted twice with water and acidified with concentrated hydrochloric acid to pH 1-2. The aqueous solution was filtered followed by extraction with chloroform. Then, a concentrated ammonium hydroxide solution was added to the aqueous solution, up to neutralization. The solution was evaporated and the obtained brown syrup was washed with methanol. Meglumine was solidified and filtered followed by silica gel addition to the methanolic solution. The solvent was evaporated and the obtained mixture was purified by silica-gel column chromatography (mobile phase: 20% methanol in dichloromethane).
This synthesis and purification affords a material of sufficient purity to be suitable for use as a standard.
The molecular structure, chemical formula and molecular weight of BH-3-HLAQ is presented below.
A 1N NaOH solution was added to a suspension of 3-HLAQ in water. The yellow solution was stirred for 0.5 hr followed by the addition of 1N HCl solution. The aqueous solution that contained white solid was extracted with ethyl acetate. The organic phase was washed with brine and dried over sodium sulfate. The suspension was filtered, evaporated to dryness and the solid residue was purified by crystallization from IPA:water mixture (1:3.75 v/v). The suspension was cooled to 0-5° C. and was kept for 1 hr, filtered and washed with IPA:water mixture (1:10 v/v). The obtained white solid was dried in a vacuum oven at 50° C.
At the time of initial qualification, the BH-3-HLAQ standard was demonstrated to conform to the molecular structure by IR, elemental analysis, MS, and NMR. Additional testing included chromatographic purity and loss on drying.
This synthesis and purification affords a material of sufficient purity to be suitable for use as a standard.
For structure elucidation purposes, a sample of BH-3-HLAQ was characterized by NMR, MS, elemental analysis and FT-IR as depicted below.
The 1H-NMR and 13C-NMR characterization of BH-3-HLAQ was performed in DMSO on a Bruker 300 MHz instrument. The peak assignments are summarized in Table 38. The 1H-NMR and 13C-NMR spectra are presented in
13C and 1H-NMR Peak Assignments for BH-3-HLAQ
13C (ppm)
1H (ppm)
The mass spectrum of BH-3-HLAQ was obtained on a Q-TOF Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray ionization in positive ion mode (ES+). The spectrum shown in
The attenuated total reflectance (ATR) FT-IR spectrum of BH-3-HLAQ was measured with a Nicolet 6700 “Thermo Scientific” FT-IR apparatus.
The test for elemental analysis was performed on a Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight percentages are accurate and reproducible to within ±0.3%. The accuracy for halogens is ±0.5%. The results for BH-3-HLAQ are presented in Table 41.
Laquinimod sodium is a white to off-white powder, freely soluble in water which should be stored in a well-closed container, protected from light, at room temperature.
The specifications of Laquinimod sodium are as follows:
Escherichia Coli - Absence/g
Table 43 lists the non-pharmacopoeial analytical methods currently in use for the determination of impurity levels in the drug substance at the time of release and during stability testing. The method numbers and the detection and quantitation limits for each impurity are also listed.
Laquinimod drug product was prepared as 0.6 mg capsules of laquinimod sodium. The specifications of laquinimod sodium (0.6 mg) capsules are as follows:
1R = Release; S = Stability
2MCQ: 5-chloro-4hydroxy-1-methylquinolin-2(1H)-one
3MCQCA: 5-chloro-4-hydroxy-1methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic acid
45-HLAQ: N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide
5MEG-LAQ: N-ethyl-4-hydroxy-1 methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxo-N-phenyl 1,2-dihydroquinoline-3-Carboxamide.
6BH-3-HLAQ: 2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)benzoic acid
7Frequency: Pilot batches, first three production batches and occasional test upon request thereafter.
8Frequency: For Release: Pilot batches, first 3 production batches and every fifth batch thereafter, or at least one batch per year if less than 5 batches per year are manufactured. For Stability: a) For long term testing at zero time and annually and/or expiry date. b) For intermediate testing at zero time and 12 months. c) For accelerated testing at zero time and 6 months.
Finished product stability protocol package sizes were the largest and smallest of each marketing container/closure and/or unidose configuration.
9For validation batches and commercial batches only.
10Only if accelerated testing fails.
The following examples describe non-pharmacopoeial analytical methods for the determination of impurity levels in the drug product at the time of release and during stability testing. The method numbers and the detection and quantitation limits for each impurity are also listed, as well as the corresponding validation numbers.
1NEA and 3-HLAQ were omitted from the commercial specifications since they were never observed. The monitoring of these impurities in the primary stability batches will continue until completion of the stability programs, using the listed method.
Laquinimod sodium drug substance was analyzed by HPLC. The amount of MCQ, MCQCA, MCQME, MCQEE and 5-HLAQ was determined using the following HPLC method.
All preparations are performed in amber flasks with non transparent caps. Immediately after preparation place the solutions in a refrigerator or in a cooled to 5° C. autosampler rack.
Weigh accurately about 15 mg of Laquinimod sodium standard into a 50 mL volumetric flask. Dilute with diluent A up to ⅔ of the volume, sonicate for about 2 minutes in cold sonication bath and dilute to volume with the Diluent A. Concentration of Stock standard solution is about 300 μg/mL. Stock standard solution may be stored for one month in refrigerator.
Dilute 3 mL of the Stock standard solution to 10 mL with diluent B (dilution factor 3.33). Concentration of Laquinimod sodium is about 90 μg/mL. Standard solution A may be used for 7 days when stored in a refrigerator.
Weigh accurately about 18 mg of MCQCA standard into a 100 mL volumetric flask. Dilute to volume with acetonitrile, sonicate until the substance is completely dissolved—stock MCQCA solution.
Concentration of MCQCA is about 180 μg/mL.
MCQCA Stock standard solution should be freshly prepared.
Prepare a solution in diluent B, containing Laquinimod in a concentration of 0.2% and MCQCA—in a concentration of 0.1%, with respect to the working concentration of Laquinimod in Standard solution A. As an example, apply the following procedure.
Transfer 4.0 mL of Laquinimod sodium standard solution for assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a 100 mL volumetric flask and dilute to volume with the diluent B (Intermediate dilution solution).
Place 2.5 mL of this intermediate dilution into a 50 mL volumetric flask and make up to volume with the diluent B.
Total dilution factor for Laquinimod standard is 1666.67, for MCQCA 2000.
Concentration of Laquinimod sodium is about 0.18 μg/mL (0.2%).
Concentration of MCQCA is about 0.09 μg/mL (0.1%, QL level).
Standard solution I may be used for 24 hours when stored in a refrigerator.
Prepare a solution of the following impurities in the Diluent A:
Mixed solution may be prepared as follows:
Weigh about 3 mg of each impurity to a 100 mL volumetric flask, dissolve and dilute to volume with the Diluent A—Mixed solution. Concentration of each impurity in the Mixed solution is about 30 μg/mL. The Mixed solution is used only for the determination of the exact retention times of the known impurities and may be used up to four months when stored at about −20° C. For this purpose, the freshly prepared Mixed solution is divided into aliquots and stored immediately at about −20° C. After thawing, the aliquots should not be refrozen.
Transfer 3 mL of the stock standard solution (Solution S) and 0.3 mL of the Mixed solution to a 10 mL volumetric flask and dilute to volume with the Diluent B. Concentration of Laquinimod sodium form is about 90 μg/mL. Concentration of each impurity is about 0.9 μg/mL (1% with respect to Laquinimod sodium). Resolution solution may be used for 9 days if stored in refrigerator.
Weigh accurately about 15 mg of Laquinimod sodium Cryst or DS into a 50 mL volumetric flask. Dilute with the Diluent A up to ⅔ of the volume, sonicate for about 2 minutes in cold sonication bath and dilute to volume with the Diluent A—Stock sample solution.
Dilute 3 mL of the Stock sample solution to 10 mL with the Diluent B—Working sample solution (dilution factor 3.33).
Concentration of Laquinimod sodium is about 90 μg/mL.
The sample must be analyzed not later than within 24 hours after preparation.
Inject the Resolution solution, Blank, Standard solutions for assay and IDD, Laquinimod sample solutions according to the relevant SOP's.
1. The typical retention time for the Laquinimod peak is 15.5±2.0 minutes.
2. The tailing factor for the Laquinimod peak should be not more than 2.0.
3. Resolution factor of NLT 2 between the peaks should be achieved.
4. The typical RRT of the specified impurity peaks should be as the following:
Evaluate Laquinimod standards for assay and IDD in order to test the system precision according to the relevant SOP's.
MCQCA in Solution I is used to test the sensitivity of the system. The RSD of the area of the six injections of Std 1 as well as the difference between Std1 and Std2 should be NMT 20%.
The RT of the main (Laquinimod) peak obtained in the sample chromatogram should correspond to that obtained for the Standard solution.
Report the calculated result of the Assay on dry basis after subtraction of Laquinimod acid content.
Calculations are performed with respect to the diluted Laquinimod sodium standard (Solution I).
Quantitation level of MCQME, MCQ, MCQEE, 5-HLAQ and unknown impurities is 0.05%.
Detection level of MCQME, MCQ, MCQEE, 5-HLAQ and unknown impurities is 0.02%.
Reporting level of MCQCA is 0.10%.
Detection level of MCQCA is 0.03%
Adjust integration parameters to reject peaks with the area less than 10% of Laquinimod sodium standard (solution I) average area.
Correlate the known impurity peaks in the sample chromatogram with those in the system suitability chromatogram within ±5% of the actual corresponding retention times.
Calculation of impurities in Laquinimod sodium:
RRF—relative response factors of impurities calculated as the following ratio: slope of Laquinimod sodium regression line/slope of impurity regression line
Values for Relative Response Factors (RRF) with respect to Laquinimod sodium are:
Report impurities as follows:
Laquinimod sodium drug substance was analyzed by HPLC. The amount of NEA, 3-HLAQ And SPIRO-LAQ was determined using the following HPLC method.
All the preparations should be performed in amber flasks. It is recommended to wash volumetric flasks and pipettes with acetonitrile before use. The use of plastic Pasteur pipettes should be excluded.
Weigh accurately about 25 μL (about 25 mg) of N-Ethylaniline (NEA) standard into a 50 mL volumetric flask. Dilute to volume with Diluent A.
Concentration of stock standard solution is about 500 μg/mL.
NEA stock standard solution may be used for three weeks when stored in a refrigerator.
Dilute 0.5 mL of NEA standard stock solution to 50 mL with Diluent B.
Concentration of N-Ethylaniline is about 5 μg/mL.
Dilute 0.5 mL of NEA standard primary dilution solution to 25 mL with Diluent B (total dilution factor—5000).
Concentration of N-Ethylaniline is about 0.1 g/mL (about 0.1% of the working concentration).
NEA working standard solution may be used for four days when stored in a refrigerator.
Weigh accurately about 15 mg of Laquinimod sodium standard into a 50 mL volumetric flask. Dilute with Diluent A up to ⅔ of the volume, sonicate for about 2 minutes in cold sonication bath and dilute to volume with Diluent A.
Laquinimod stock solution may be used for one month when stored in refrigerator.
Weight about 2.5 mg of SPIRO-LAQ into a 50 mL volumetric flask. Dissolve and dilute to volume with a mixture of 95% acetonitrile—5% water.
Weight about 2.5 mg of 3-HLAQ into a 50 mL volumetric flask. Dissolve and dilute to volume with 95% acetonitrile—5% water.
Transfer the following aliquots to 50 mL volumetric flask and dilute to volume with Diluent B:
15 mL of Laquinimod sodium stock solution, 1.0 mL of SPIRO-LAQ stock solution, 1.0 mL of 3-HLAQ stock solution and 0.1 mL of NEA standard stock solution.
Concentration of Laquinimod sodium is about 90 μg/mL.
Concentration of NEA, SPIRO-LAQ and 3-HLAQ is about 1 μg/mL (1%).
The Resolution Solution is used only for the determination of the retention times of Laquinimod, NEA, 3-HLAQ and SPIRO-LAQ and may be used for 38 days when stored at −20° C. For this purpose, the freshly prepared resolution solution should be divided into aliquots and stored immediately at −20° C. After thawing, the solution aliquots should not be refrozen.
Mix vigorously resolution solution after thawing before injection.
After thawing, the Resolution Solution may be used for 4 days when stored in refrigerator.
Weigh accurately about 15 mg of Laquinimod sodium Cryst or DS into a 50 mL volumetric flask. Dilute with Diluent B up to ⅔ of the volume, sonicate until the substance is completely dissolved and dilute to volume with Diluent B—sample stock solution.
Dilute 3 mL of sample stock solution to 10 mL with Diluent B (dilution factor 3.33).
Concentration of Laquinimod sodium is about 90 μg/mL.
Note: Immediately after preparation put the sample solution to refrigerator or at cooled to 5° C. auto sampler. The sample must be analyzed not later than 12 hours after preparation.
Inject resolution solution, Diluent B (two or three times), Standard and Sample solutions according to relevant SOP.
The typical retention time is 2.2±0.5 minutes for Laquinimod peak.
The typical retention time is 12.2±1.5 minutes for NEA peak.
The chromatographic mapping of the specified impurity peaks should be in the following order: 3-HLAQ, NEA and SPIRO-LAQ.
Resolution of NLT 2 for both pairs of peaks should be achieved.
Evaluate standards in order to test system precision according to the relevant SOP.
Inject Diluent B to detect system peaks.
Quantitation limit for NEA and any other impurities is 0.06%
Detection limit for NEA and any other impurities is 0.02%.
Adjust integration parameters to reject peaks with the area less than 15% of standard area.
Disregard all the impurity peaks eluting between 0 and the RT of Laquinimod plus one minute.
Calculate the content of non-polar impurities following the equation:
Where RRF is the Relative Response Factor of NEA to impurities as per the Table:
Laquinimod capsules containing 0.6 mg Laquinimod were analyzed by HPLC. The amount of MCQ, MCQCA, 5-HLAQ, MCQME and MCQEE was determined using the following HPLC method.
All the preparations should be protected from light. For liquid preparations use amber flasks with non transparent caps. Use only glass Pasteur pipettes. Standard and sample solutions should be kept refrigerated at 2-8° C. after preparation.
Weigh accurately in duplicate about 15 mg of Laquinimod sodium standard into a 50 mL volumetric flask. Dilute with diluent A up to ⅔ of the volume, sonicate for 2 minutes in a cold sonication bath and dilute to volume with diluent A.
Concentration of standard stock solution is about 300 μg/mL Laquinimod sodium.
Standard stock solution may be used for one month when stored in a refrigerator (2° C.-8° C.).
Dilute 3 mL of the Standard Stock Solution to 10 mL with diluent B (Dilution factor 3.33).
Concentration of Laquinimod sodium is about 90 μg/mL. Concentration expressed as Laquinimod (acid) is about 85 μg/mL.
Standard working solution A may be used for 7 days when stored in a refrigerator (2° C.-8° C.).
Weigh accurately about 18 mg of MCQCA standard into a 100 mL volumetric flask. Dilute to volume with acetonitrile, sonicate (in a cold sonication bath) until the substance is completely dissolved—stock MCQCA solution.
Concentration of MCQCA is about 180 μg/mL.
MCQCA Stock standard solution should be freshly prepared.
Prepare a solution in diluent B, containing Laquinimod in a concentration of 0.2% and MCQCA—in a concentration of 0.1%, with respect to the working concentration of Laquinimod in Standard solution A. As an example, apply the following procedure.
Transfer 4.0 mL of Laquinimod sodium standard solution for assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a 100 mL volumetric flask and dilute to volume with the diluent B (Intermediate dilution).
Place 2.5 mL of this intermediate dilution into a 50 mL volumetric flask and make up to volume with the diluent B.
Total dilution factor for Laquinimod standard is 1666.67, for MCQCA 2000.
Concentration of Laquinimod sodium is about 0.18 μg/mL (0.2%).
Concentration of MCQCA is about 0.09 μg/mL (0.1%, QL level).
Standard solution I may be used for 24 hours when stored in a refrigerator.
Prepare solution containing the following potential impurities standards (markers) using the Diluent A as a solvent:
The Mixed Solution may be prepared as follows:
Weigh about 3 mg of each impurity standard/marker into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A.
Concentration of each impurity in the Mixed Solutions is about 30 μg/mL.
Mixed Solution may be used for up to 4 months when stored frozen at about −20° C.
For this purpose, the freshly prepared Mixed Solution should be divided into aliquots, immediately frozen and stored at −20° C. After thawing, the aliquots should be mixed well and should not be refrozen.
Weigh about 3 mg of MCQEE (Ethyl 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate) into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A. This is MCQEE Stock solution.
This Solution may be used for up to 4 months when stored frozen at about −20° C.
For this purpose, the freshly prepared MCQEE Stock Solution should be divided into aliquots, immediately frozen and stored at −20° C. After thawing, the aliquots should be mixed well and should not be refrozen.
Weigh about 3 mg of MEG-LAQ (Meglumine Adduct of Laquinimod) into a 100 mL volumetric flask, dissolve (sonication is acceptable) and dilute to volume with the Diluent A. This is MEG-LAQ Stock solution.
This Solution may be used for one week when stored in a refrigerator (2° C.-8° C.).
Prepare two Resolution Solutions separately as follows, using the Mixed solution.
Transfer 3 mL of Laquinimod standard stock solution (Solution S), 0.3 mL of the Mixed solution and 0.3 mL of the MCQEE Stock solution to a 10 mL volumetric flask and dilute to volume with the Diluent B. This is Resolution Solution 1.
Concentration of Laquinimod sodium in it is about 90 μg/mL. Concentration of each impurity—is about 0.9 g/mL (about 1% with respect to the working concentration of Laquinimod).
Resolution Solution 1 is used for resolution test (for system suitability) and for determination of retention times (RT)/relative retention times (RRT) of five known impurities: MCQ, MCQCA, MCQME, MCQEE and 5-HLAQ.
Resolution Solution 1 may be used for 9 days if stored in a refrigerator (2° C.-8° C.).
Note: Instead of Resolution Solution i, a Resolution Solution prepared as per the Method of Example 19 may be used.
Transfer 3 mL of Laquinimod standard stock solution (Solution S), 0.3 mL of the Mixed solution and 0.3 mL of the MEG-LAQ Stock solution to a 10 mL volumetric flask and dilute to volume with the Diluent B. This is Resolution Solution 2.
Concentration of Laquinimod sodium in it is about 90 g/mL. Concentration of each impurity—is about 0.9 μg/mL (about 1% with respect to the working concentration of Laquinimod).
Resolution Solution 2 is used for determination of retention time of MEG-LAQ.
Resolution Solution 2 may be used for one week if stored in a refrigerator (2° C.-8° C.).
Weigh accurately 20 capsules and completely empty their contents into a mortar. Pay attention to complete emptying of capsule contents into the mortar, using spatula when necessary. Weigh the empty capsules. Calculate the average weight of the capsule contents.
Mix and grind the capsule contents in a mortar and keep the powder in a tightly closed container protected from light.
Weigh accurately, in duplicate, the amount of powder corresponding to 7 capsules, into a 50 mL volumetric flask.
Add diluent B up to ⅔ of the volume, shake for 30 minutes at 200 mot/min. Dilute to volume with diluent B. Mix well. Filter before use through a 0.45 μm GHP ACRODISC GF filter or equivalent, discarding the first 0.5-1 mL.
Working concentration of Laquinimod (acid) is about 84 μg/mL.
Note: Immediately after preparation place sample solutions into a refrigerator or in a cooled to 5° C. autosampler. The sample solutions may be used for 24 hours when kept at the temperature 2° C.-8° C.
Inject the Resolution Solutions, Diluent B (Blank), Standard Solutions for assay and IDD and Sample solutions, according to the relevant SOP's.
Determine the retention time (RT) and the area of the Laquinimod peak in the chromatograms of Sample and Standard Solutions for identification and assay.
Determine the RT, the relate rive retention time (RRT) and peak areas of all known impurities and any other impurities in the chromatograms of Sample Solutions, for calculation of the content of impurities/degradation products.
Ignore early eluting peaks of excipients and system peaks (See chromatogram for determination of impurities/degradation products). For example, use integration inhibition between 0 and RRT 0.15 (about 2.5 minutes).
Adjust integration parameters to reject peaks with area less than 10% of the average response of Laquinimod peak in the injections of Standard Solution I (for determination of impurities/degradation products).
Disregard peak of MEG-LAQ in sample injections (identified following Resolution Solution 2). The content of MEG-LAQ will be tested by a different method.
Typical retention time of Laquinimod peak is 15.5±2.0 minutes.
Tailing factor (USP) for Laquinimod peak should be not more than 2.0.
Resolution factor for all the pairs of peaks should be NLT 2.
RRT of the peaks of known impurities/degradation products should be as follows:
MEG-LAQ peak is substantially broadened in comparison with neighboring peaks. Retention time of MEG-LAQ is variable, being very sensitive to slightest changes in chromatographic conditions (pH, % Acetonitrile, Temperature, etc.) and therefore should be defined using its peak in the chromatogram of the Resolution Solution 2. Typically, its RRT is about 0.66.
Evaluate Laquinimod standards for assay and IDD in order to test the system precision according to the relevant SOP's.
MCQCA in Solution I is used to test the sensitivity of the system. RSD of the area of six injections of Std 1 as well as the difference between Std 1 and Std 2 should be NMT 20%.
Inject diluent B to detect system peaks.
The RT of the main peak obtained in the sample chromatogram should correspond to that obtained for the Laquinimod peak in the injection of Standard Solution.
Where: 0.94: Conversion factor of Laquinimod sodium salt to Laquinimod (acid).
AreaImpurity: Area of an impurity/degradation product (known or unknown) peak in Sample Solution.
Areastd: Area of Laquinimod peak in chromatogram of Standard Solution I.
0.94: Conversion factor of Laquinimod sodium salt to Laquinimod (acid).
RRF: Relative response factors of impurities/degradation products calculated as the following ratio: slope of Laquinimod regression line/slope of impurity regression line.
The values for relative response factors with respect to Laquinimod are:
RRF for unknown impurities/degradation products is taken as 1.0.
Quantitation level (QL) of MCQME, MCQ, MCQEE, 5-HLAQ and unknown impurities is 0.05%. Detection level (DL) of MCQME, MCQ, MCQEE, 5-HLAQ and unknown impurities is 0.02%.
QL of MCQCA is 0.1%. Detection level DL of MCQCA is 0.03%.
Correlate all the peaks in sample chromatogram with those in the system suitability chromatogram, within ±5% of the actual corresponding retention times.
Report as follows:
Laquinimod capsules containing 0.6 mg Laquinimod were analyzed by HPLC. The amount of NEA, 3-HLAQ And SPIRO-LAQ was determined using the following HPLC method.
All the preparations should be protected from light. For liquid preparations use amber flasks with non transparent caps.
Use only glass Pasteur pipettes.
Standard and sample solutions should be kept refrigerated at 2-8° C. after preparation.
Weigh accurately in duplicate about 18 μL (about 18 mg) of N-Ethylaniline (NEA) standard into a 50 mL volumetric flask. Dilute to volume with acetonitrile.
Concentration of standard stock solution is about 360 μg/mL.
NEA standard stock solution may be used for three weeks when stored in a refrigerator (2° C.-8° C.).
Dilute 0.5 mL NEA standard stock solution to 50 mL with acetonitrile.
Concentration of standard primary dilution solution is about 3.6 μg/mL.
Transfer 0.5 mL NEA standard primary dilution solution to a 20 mL volumetric flask. Add 4 mL diluent A and dilute to volume with acetonitrile.
Concentration of NEA is about 0.09 μg/mL (about 0.1% of the Laquinimod working concentration).
NEA working standard solution may be used for one week when stored in a refrigerator (2° C.-8° C.).
Resolution solution from Example 20 may be used.
Weigh accurately about 15 mg of Laquinimod sodium standard into a 50 mL volumetric flask. Dilute with acetonitrile:water [50:50 (v/v)] up to ⅔ of the volume, sonicate for 2 minutes in a cold sonication bath and dilute to volume with acetonitrile:water [50:50 (v/v)].
Concentration of Laquinimod stock solution is about 300 μg/mL.
Laquinimod stock solution may be used for one month when stored in a refrigerator (2° C.-8° C.).
Weigh accurately about 25 μL (about 25 mg) of N-Ethylaniline (NEA) standard into a 50 mL volumetric flask. Dilute to volume with acetonitrile:water [50:50 (v/v)].
Concentration of NEA stock solution is about 500 μg/mL.
NEA stock solution may be used for 3 weeks if stored in a refrigerator (2° C.-8° C.).
Weigh about 2.5 mg of SPIRO-LAQ into a 50 mL volumetric flask. Dissolve and dilute to volume with acetonitrile:water [95:5 (v/v)].
Concentration of SPIRO-LAQ stock solution is about 50 g/mL.
Weigh about 2.5 mg of 3-HLAQ into a 50 mL volumetric flask. Dissolve and dilute to volume with acetonitrile:water [95:5 (v/v)].
Concentration of 3-HLAQ stock solution is about 50 μg/mL
Transfer the following aliquots to a 50 mL volumetric flask and dilute to volume with mobile phase solution B [acetonitrile:buffer pH 7.0 (65:35 v/v)]:
15 mL of Laquinimod stock solution for resolution test,
1.0 mL of SPIRO-LAQ stock solution for resolution test,
1.0 mL of 3-HLAQ stock solution for resolution test,
0.1 mL of NEA stock solution for resolution test.
Concentration of Laquinimod sodium is about 90 μg/mL.
Concentration of NEA, SPIRO-LAQ and 3-HLAQ is about 1 μg/mL (about 1% with respect to Laquinimod working concentration).
The resolution solution is used only for the determination of the retention times of the laquinimod and known potential impurities/degradation products and may be used for 38 days when stored at −20° C. For this purpose, the freshly prepared resolution solution should be divided into aliquots and stored immediately at −20° C. After thawing, the solution aliquots should be mixed vigorously and should not be refrozen.
The resolution solution may be used for 4 days when stored in a refrigerator (2° C.-8° C.).
Weigh accurately at least 10 capsules and empty the contents in a mortar. Weigh the empty capsules. Calculate the average weight of the capsule contents.
Grind the sample in a mortar and keep the powder in a tightly closed protected from light container.
Weigh accurately, in duplicate, the amount of powder corresponding to 3 Laquinimod capsules into a 20 mL volumetric flask.
Add 4 to 10 mL of Diluent A and shake for 15 minutes at 200 mot/min to dissolve. Dilute to volume with acetonitrile. Mix well. Filter before use through a 0.45 μm GHP Acrodisc GF filter or equivalent, discarding the first 2 mL.
Concentration of Laquinimod is about 90 μg/mL.
Note: Immediately after preparation place the sample solution into a refrigerator or in a cooled to 5° C. autosampler. The sample should be analyzed not later than within 24 hours after preparation.
Inject resolution solution, blank, standard and sample solutions according to the relevant SOP's for impurities/degradation products determination.
Determine the peak area of NEA in standard working solutions.
Determine the retention time (RT) of the Laquinimod peak in the chromatograms from resolution and sample solutions
Determine the RT, the relative retention time (RRT) to Laquinimod peak and the peak area of all known impurities and unknown impurities in the chromatograms from Sample solutions.
Apply integration inhibition between 0 and the 1.6×RT of Laquinimod peak (RT of system peak).
Adjust integration parameters to reject peaks with the area less than 20% of NEA standard area.
The typical retention time for Laquinimod is 2.2±0.5 minutes.
The typical retention time for NEA is 12.2±1.5 minutes.
The RRT of the specified impurity/degradation product peaks relative to NEA should be as follows:
Resolution factor of NLT 2 for both known impurities/degradation products peaks from NEA peak should be achieved.
Evaluate standards in order to test the system precision according to the relevant SOP for impurities/degradation products determination.
Inject blank to detect system peaks.
Calculate % IDD in Laquinimod capsules as follows:
AreaImp: Area of the NEA, 3-HLAQ or unknown impurity/degradation product peak in the sample solution.
See note in 8.3.
AreaStd NEA: Area of the NEA in standard working solution.
RRF: Relative response factors of impurities calculated as the following ratio:
Slope of NEA regression line/slope of impurity regression line.
Note: For Unknown impurities/degradation product, the slope of Laquinimod regression line was used.
The values for relative response factors with respect to NEA are:
Quantitation limit (QL) for NEA, 3-HLAQ and any other unknown impurity/degradation product is 0.1% with respect to the Laquinimod working concentration.
Detection limit (DL) for NEA, 3-HLAQ and any other unknown impurity/degradation product is 0.03% with respect to the Laquinimod working concentration.
Report content of Impurities/Degradation products as follows:
Note: Peaks eluting at RT of the SPIRO-LAQ (±5% of the actual retention time in system suitability test) report as “any other impurity” peak.
Laquinimod capsules containing 0.6 mg Laquinimod were analyzed by HPLC. The amount of MEG-LAQ and BH-3-HLAQ was determined using the following HPLC method.
Mobile phase parameters, flow rate and column temperature may be altered in order to achieve the system suitability requirements. The Mobile phase parameters may be altered in the range: 55%-62% (v/v) Solvent A and 45%-38% (v/v) Solvent B. All the solvents must be of HPLC grade or equivalent. All the preparations should be kept refrigerated at 2-8° C. and be protected from light. Use amber flasks (low actinic glassware) with non transparent caps. Avoid heating of sonication bath.
Prepare in duplicate. Accurately weigh about 11.5 mg of BH-3-HLAQ into a 50 mL volumetric flask, sonicate to dissolve and dilute to volume with the Diluent 1.
Concentration of BH-3-HLAQ stock solution is about 230 μg/mL.
BH-3-HLAQ stock solution may be used for 1 week if stored refrigerated (2° C.-8° C.).
Prepare in duplicate. Accurately weigh about 11.5 mg of MEG-LAQ into a 50 mL volumetric flask, sonicate to dissolve and dilute to volume with the Diluent 1. Concentration of MEG-LAQ stock solution is about 230 μg/mL.
MEG-LAQ stock solution may be used for 1 week if stored refrigerated (2° C.-8° C.).
Note: Due to high hygroscopicity of MEG-LAQ, its exposure to air should be minimal: close the vial with standard immediately after weighing; it is recommended to weigh MEG-LAQ under controlled low humidity conditions.
Transfer 2.0 mL of BH-3-HLAQ Stock solution to a 50 mL volumetric flask and dilute to volume with Diluent 2. Concentration of BH-3-HLAQ is about 9.2 μg/mL.
Transfer 2.0 mL of MEG-LAQ Stock solution to a 50 mL volumetric flask and dilute to volume with Diluent 2. Concentration of MEG-LAQ is about 9.2 μg/mL.
Transfer 1.0 mL of the Primary Diluted Solution of BH-3-HLAQ and 2.5 mL of the Primary Diluted Solution of MEG LAQ to a 50 mL volumetric flask and dilute to volume with Diluent 2.
Total dilution factor in this standard solution is: 1250 for BH-3-HLAQ and 500 for MEG-LAQ (from the stock solution).
Concentration of BH-3-HLAQ is about 0.18 μg/mL (0.2% of Laquinimod working concentration), and of MEG-LAQ is about 0.45 μg/mL (0.5% of Laquinimod working concentration).
Standard solution may be used for 1 week if stored refrigerated (2° C.-8° C.).
Prepare once. Transfer 0.5 mL of Primary Diluted Solution of BH-3-HLAQ and 2.0 mL of the Primary Diluted Solution of MEG LAQ (one of the duplicates each) to a 100 mL volumetric flask and dilute to volume with Diluent 2.
Concentration of BH-3-HLAQ is about 0.045 μg/mL (0.05% of Laquinimod working concentration), and of MEG-LAQ is about 0.18 g/mL (0.2% of Laquinimod working concentration).
Prepare once. Weigh about 15 mg of Laquinimod sodium standard into a 50 mL volumetric flask. Dilute with the Diluent 1 up to ⅔ of the volume, sonicate for 2 minutes in a sonication bath and dilute to volume with the Diluent 1. Concentration of Laquinimod stock solution is about 300 μg/mL Laquinimod sodium.
Laquinimod Stock solution may be used for 1 month if stored refrigerated (2° C.-8° C.).
Transfer 3 mL of Laquinimod stock solution and 1 mL of each of the Primary Diluted solutions (one of the duplicates of each) to a 10 mL volumetric flask. Dilute to volume with Diluent 2. Mix well.
Concentration of Laquinimod Sodium in the Resolution Solution is about 90 μg/mL (about 85 g/mL if expressed as Laquinimod—free acid). Concentration of each impurity is about 0.9 μg/mL (about 1% with respect to the working concentration of Laquinimod).
Resolution Solution may be used for one week if stored refrigerated (2° C. 8° C.).
Weigh accurately 20 capsules and completely empty their contents into a mortar. Pay attention to complete emptying of capsule contents into the mortar, using spatula when necessary. Weigh the empty capsules. Calculate the average weight of the capsule contents.
Mix and grind the capsule contents in a mortar and keep the powder in a tightly closed container protected from light.
Weigh accurately, in duplicate, the amount of powder corresponding to 7 capsules, each into a separate 50 mL volumetric flask.
Add diluent 2 up to ⅔ of the volume, shake for 30 minutes at 200 mot/min. Dilute to volume with diluent 2. Mix well. Filter before use through a 0.45 μm ACRODISC GHP GF filter or equivalent, discarding the first 0.5-1 mL.
Immediately after preparation place sample solutions into a refrigerator or in a cooled to 5° C. autosampler.
Working concentration of Laquinimod (acid) is about 84 μg/mL.
Sample solutions may be used for 24 hours when kept at the temperature 2° C.-8° C.
Inject the Resolution solution once.
Typical retention time of Laquinimod peak* is within the range 12-18 minutes.
Order of elution of peaks in the injection of Resolution Solution and typical RRT of the impurities peaks, with respect to Laquinimod peak, are as follows:
Flow rate, temperature and mobile phase composition (ratio of Solvent A and Solvent B) may be adjusted to achieve the required system suitability parameters.
Inject Blank (Diluent 2) twice to detect system peaks. Disregard system peaks.
From 6 replicate injections of the QL Solution, evaluate areas of BH-3-HLAQ and MEG-LAQ peaks.
RSD for each of the peaks, in 6 replicates, should be not more than 20%.
Evaluate areas of BH-3-HLAQ and MEG-LAQ peaks in the chromatograms of the standard injections in order to test the system precision according to relevant SOP's.
Inject Resolution Solution, Blank—twice, QL solution (6 replicates), Standard solutions and Sample solutions, according to relevant SOP's.
Disregard all the peaks other than MEG-LAQ and BH-3-HLAQ in the injections of standard and sample solutions.
Determine the retention time (RT) and the areas of the peaks of MEG-LAQ and BH-3-HLAQ in the chromatograms of Sample solutions, for identification and quantitation.
Adjust integration parameters to reject peaks with area less than 10% of the average response of BH-3-HLAQ peak in the chromatograms of six successive injections of the first preparation of Standard Solution.
This corresponds to 0.02% (DL of BH-3-HLAQ) with respect to working concentration of Laquinimod.
Quantitation of each of the impurities (MEG-LAQ and BH-3-HLAQ) is performed with respect to the corresponding peaks in the chromatograms of the Standard solutions.
Use the following equation to calculate the content of MEG-LAQ and BH-3-HLAQ:
AreaSmp: Area of MEG-LAQ or BH-3-HLAQ peak in the chromatograms of the Sample solutions
AreaStd: Area of peak MEG-LAQ (average) or BH-3-HLAQ peak (average), respectively, in the chromatograms of Standard solutions
Label Claim: 0.6 mg
QL (Quantitation limit) of BH-3-HLAQ is 0.05%; QL of MEG-LAQ is 0.2%
DL (Detection limit) of BH-3-HLAQ is 0.02%; DL of MEG-LAQ is 0.1%
For the values equal or above QL, report the calculated result.
For the values below QL but above DL, report: “Less than 0.05% (QL)” for BH-3-HLAQ and “Less than 0.2% (QL)” for MEG-LAQ.
For the values below DL, report: “Less than 0.02% (DL)” for BH-3-HLAQ and “Less than 0.1% (DL)” for MEG-LAQ.
DEREK prediction software (DEREK Nexus, version 2.0.0, 2011i, LHASA Ltd, Leeds, UK) was used to evaluate the genotoxic potential of MCQ and MCQME. Computational toxicology evaluation is summarized in Table 47.
The DEREK definition of the likelihood level of a structure being toxic is as follows:
Plausible: The weight of evidence supports the proposition.
Equivocal: There is an equal weight of evidence for and against the proposition.
A number of batches of laquinimod sodium drug substance were manufactured at various manufacturing facilities and subsequently analyzed. During manufacture and analysis, sufficient light protection of sample solution was provided.
All batches of drug substance were within specification as provided in Example 17.
Table 48 and Table 49 shows batch analyses of laquinimod sodium drug substance batches Table 50 shows batch analyses of laquinimod acid.
0.05%1
1Reporting limit 0.10%.
4Deviation from method. Different mobile phase and wavelength used.
Impurities were observed in laquinimod sodium batches at release and after long-term and accelerated storage.
The ranges of impurity levels observed in the laquinimod sodium batches, including the primary stability and validation batches, are summarized in Table 51.
A number of batches of laquinimod sodium drug product were manufactured at various manufacturing facilities and subsequently analyzed. During manufacture and analysis, sufficient light protection of sample solution was provided.
All batches of drug product were within specification.
Evaluation of the release data shows that all batches have been within the specifications in effect at the time of their release. Furthermore it can be seen that all manufactured batches of the product proposed for commercialization, e.g., laquinimod 0.6 mg capsules, have shown compliance with the proposed commercial specifications. Batch-to-batch consistency over time was found in the results for all the measured parameters. The batch analysis results demonstrate that the quality of all batches does not differ significantly from each other and that production at different manufacturing sites affords material of similar quality.
Table 52, Table 53, Table 54, Table 55, Table 55, Table 56 and Table 57 relate to batch analyses of laquinimod sodium drug product batches.
4The analysis was performed using the previous method for determination of MEG-LAQ.
5The test for BH-3-HLAQ was introduced after the release of these batches. The listed results
6For batch 27 in Alu/Alu blisters a result of 0.06% was obtained.
1NEA and 3-HLAQ were omitted from the commercial specifications since they were never
6Determination of BH-3-HLAQ and water content were introduced in the specifications
1NEA and 3-HLAQ were omitted from the commercial specifications since they were
4An additional specified impurities were reported: 0.03% of MCQME.
This application claims priority of U.S. Provisional Application No. 61/785,575, filed Mar. 14, 2013, the entire content of which is hereby incorporated by reference herein. Throughout this application, various publications are referred to by first author and year of publication. Full citations for these publications are presented in a References section immediately before the claims. Disclosures of the publications cited in the References section in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art as of the date of the invention described herein.
Filing Document | Filing Date | Country | Kind |
---|---|---|---|
PCT/US2014/029292 | 3/14/2014 | WO | 00 |