A method for preparing impurities degraded by lentilana

Abstract

This invention discloses a method for preparing lentilana-degraded impurities. Starting with 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid and glycine ester, a condensation reaction is carried out under a condensing agent catalysis, followed by ester hydrolysis under alkaline conditions to obtain lentilana-degraded impurities. This invention features a concise synthetic route, mild reaction conditions, simple operation, and a purity exceeding 96.0%, avoiding the problem of low purification efficiency associated with chromatographic column separation.

Description

Technical Field

[0001] This invention belongs to the field of chemistry or medicinal chemistry, and specifically relates to a method for preparing lentilana to degrade impurities. Background Technology

[0002] Lotilanar (also known as Lotilanar) is a new generation of isoxazoline insecticide developed by Eli Lilly and Company. Its chemical name is 5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-3-methyl-thiophene-2-carboxylic acid (2,2,2-trifluoroethylamino-formyl)-formamide. It has a pair of enantiomers; the S-configuration has 100 times the insecticidal activity of the R-configuration and exhibits no cross-resistance with dieldrin and fipronil. In April 2017, the European Medicines Agency approved its marketing for the treatment of flea and tick infestations in dogs and cats. In December 2019, the FDA approved its marketing for the treatment and prevention of flea infestations in cats.

[0003] Lotilaner eye drops are an antiparasitic eye drop used to treat Demodex blepharitis in adults 18 years of age and older. Lotilaner inhibits the γ-aminobutyric acid (GABA)-gated chloride channels in Demodex mites, leading to scarring and death of the mites, and relieving eyelid redness and crusting. In July 2023, XDEMVY (lotilaner eye drops 0.25%), developed by Tarsus Pharmaceuticals, was the first drug approved in the United States for the treatment of Demodex blepharitis. This drug is the first and currently the only FDA-approved treatment directly targeting Demodex mites. The chemical structure of lentilana is as follows:

[0004]

[0005] Because lentilana contains amide bonds, it can undergo amide hydrolysis under acidic or alkaline conditions. The C=O bond and -NH2 group in the amide molecule react with water to generate the corresponding acid and amine. Since lentilana contains two amide bonds, there are two hydrolysis pathways. The degradation pathways and degradation impurities are confirmed by MS and are shown in the following figure:

[0006]

[0007] S-isoxazolethiophene carboxylic acid degradation impurities

[0008] Research on degradation impurities is crucial for drug quality control, but purchasing these impurities is expensive, and separation via preparative columns is inefficient, time-consuming, and yields small quantities. One of the two degradation impurities for lentilana is S-isoxazolethiophenic acid, an intermediate in the n-1 step of lentilana synthesis, which only requires purification. However, the other degradation impurity requires the development of a chemical synthesis method for the batch synthesis of high-purity degradation impurities. Summary of the Invention

[0009] The purpose of this invention is to provide a method for preparing high-purity lentilana by degrading impurities, which features a short synthetic route, mild reaction conditions, simple operation, and the following specific technical solution:

[0010] A method for preparing lentilana to degrade impurities, characterized by preparation via the following chemical reaction equation:

[0011]

[0012] Wherein, R1 = any one of methyl, ethyl, propyl, and isopropyl.

[0013] The specific preparation steps for the above reaction equation are as follows:

[0014] (1) Condensation: In a solvent, 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophen-2-carboxylic acid, condensing agent, and glycine ester are added in a certain proportion. The mixture is heated and kept at the temperature. After the temperature is maintained, water is added to separate the layers. The organic layer is concentrated under reduced pressure to dryness to obtain the condensate.

[0015] (2) Hydrolysis: The condensate is dissolved in a hydrolysis solvent, alkali is added in a certain proportion, the temperature is raised, the reaction is kept at the temperature, the temperature is lowered, acid is added to adjust the pH value to acidic, the mixture is filtered, washed with water to neutral, and dried to obtain lentilana degraded impurities.

[0016] The solvent is at least one selected from ethyl acetate, isopropyl acetate, butyl acetate, pentyl acetate, isoamyl acetate, and hexyl acetate, and the amount used is 2-15 times the weight of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid; the condensing agent is at least one selected from dicyclohexylcarbodiimide, N,N-carbodiimidazole, diisopropylcarbodiimide, 1-hydroxybenzotriazole, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, and the amount used is 2-15 times the weight of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid; The molar ratio of isoxazol-3-yl]-thiophen-2-carboxylic acid to the feed is 0.3-1.5:1; the glycine ester is at least one of glycine methyl ester, glycine ethyl ester, glycine propyl ester, and glycine isopropyl ester, and the molar ratio of its dosage to that of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisooxazol-3-yl]-thiophen-2-carboxylic acid is 1.05-2:1; the reaction temperature is 25-40℃; the reaction time is 3-7 hours; the amount of water added is 0.5-2 times the amount of solvent; the vacuum concentration temperature is 30-80℃, the vacuum degree is -0.06~-0.1MPa, and the endpoint of vacuum distillation is no continuous distillate out.

[0017] The hydrolysis solvent is at least one of methanol, ethanol, isopropanol, and acetone, and its amount is 2-10 times the weight of the condensate; the alkali is at least one of ammonia, triethylamine, potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, and lithium hydroxide, and its molar ratio to the cyclized compound is 1.5-3:1; the heat preservation reaction temperature is 50-80℃; the heat preservation reaction time is 2-6 hours; the cooling endpoint temperature is 10-30℃, and the cooling time is 0.5-3 hours; the pH adjustment endpoint is 1-3, and the acid used is at least one of phosphoric acid, sulfuric acid, hydrochloric acid, nitric acid, phosphoric acid, formic acid, acetic acid, and propionic acid; the drying temperature is 50-80℃, and the drying time is 10-16 hours.

[0018] The beneficial effects of this invention are as follows:

[0019] 1) The method for preparing lentilana to degrade impurities provided by this invention has a short synthetic route. The starting material 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid is an intermediate for the preparation of lentilana, and the starting material is readily available. The reaction conditions are mild, the operation is simple, and the yield is high, with an overall yield of 75%.

[0020] 2) According to the preparation method provided by the present invention, the content of the degradation impurities prepared can reach more than 96.0%, which can be used as a reference standard for lentilana degradation impurities for quality research and quality control. Attached Figure Description

[0021] Figure 1 Mass spectra of impurities degraded by lentilana

[0022] Figure 2 1H NMR spectrum of impurities degraded by lentilana

[0023] Figure 3 Carbon NMR spectrum of impurities degraded by lentilana Detailed Implementation

[0024] The present invention will be described in detail below through specific embodiments. These embodiments are used to explain the present invention, but not to limit it.

[0025] Example 1

[0026] (1) Condensation: 45.8 g (0.1 mol) of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid and 300 g of isoamyl acetate were added to a 1 L reaction flask. 13.4 g (0.13 mol) of glycine ethyl ester and 21.1 g (0.13 mol) of N,N-carbonyldiimidazole were added under stirring. The mixture was kept at 30 °C for 7 hours. After the reaction was completed, 150 g of water was added and stirred for 20 minutes. The mixture was allowed to stand and separate into layers. The upper organic layer was concentrated under reduced pressure until no continuous distillate flowed out. The temperature was 60 °C and the vacuum degree was -0.09 MPa, yielding a reddish-brown viscous condensate.

[0027] (2) Hydrolysis: Add the above condensate (0.1 mol), 320 g of methanol, and 6.0 g (0.25 mol) of lithium hydroxide to a 500 ml reaction flask. Heat to 60-65 °C and maintain for 6 hours, then cool to 10-15 °C within 1 hour. Adjust the pH to 2-3 with phosphoric acid. Filter and wash with water until neutral. Dry at 50 °C for 16 hours to obtain 39.4 g of lentilana degradation impurities. The two-step yield was 76.5%. The HPLC purity was 96.2%, and the condensate content was 1.5%.

[0028] MS(m / z): [MH] - = 514.94 (See details) Figure 1 )

[0029] 1HNMR (600 MHz, CD3OD): δ 7.75 (s, 2H), 7.25 (s, 1H), 4.26 (s, 1H), 3.99 (s, 1H), 3.93 (d, 2H), 2.48 (s, 3H). (See details) Figure 2 )

[0030] 13 CNMR (600 MHz, CD3OD): δ178.60, 166.99, 156.68, 144.72, 140.14, 138.19-138.31, 136.27, 133.70, 131.07, 128.63, 126.75, 90.95-91.15, 46.88-47.22, 40.98, 39.48, 38.06, 23.82, 18.14. (See details) Figure 3 )

[0031] Example 2

[0032] (1) Condensation: 45.8 g (0.1 mol) of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophene-2-carboxylic acid and 450 g of isopropyl acetate were added to a 1 L reaction flask. 10.7 g (0.12 mol) of glycine methyl ester and 26.8 g (0.13 mol) of dicyclohexylcarbodiimide were added under stirring. The mixture was kept at 35 °C for 4 hours. After the reaction was completed, 300 g of water was added and stirred for 20 minutes. The mixture was allowed to stand and separate into layers. The upper organic layer was concentrated under reduced pressure until no continuous distillate flowed out. The temperature was 60 °C and the vacuum degree was -0.09 MPa to obtain a reddish-brown viscous condensate.

[0033] (2) Hydrolysis: Add the above condensate (0.1 mol), 150 g of methanol, and 40 g (0.3 mol) of 30% sodium hydroxide solution to a 500 ml reaction flask. Heat to 65-70 °C and maintain the temperature for 5 hours, then cool to 10-15 °C within 2 hours. Adjust the pH to 2-3 with hydrochloric acid. Filter and rinse with water until neutral. Dry at 75 °C for 10 hours to obtain 39.0 g of lentilana degradation impurities. The two-step yield was 75.8%. The HPLC purity was 97.0%, and the condensate content was 1.2%.

[0034] Example 3

[0035] (1) Condensation: 45.8 g (0.1 mol) of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophen-2-carboxylic acid and 91.6 g of ethyl acetate were added to a 1 L reaction flask. 17.81 g (0.2 mol) of glycine methyl ester was added under stirring. The mixture was kept at 25 °C for 3 hours. After the reaction was completed, 50 g of water was added and stirred for 30 minutes. The mixture was allowed to stand and separate into layers. The upper organic layer was concentrated under reduced pressure until no continuous distillate flowed out. The temperature was 40 °C and the vacuum degree was -0.09 MPa to obtain a reddish-brown viscous condensate.

[0036] (2) Hydrolysis: Add the above condensate (0.1 mol), 200 g of ethanol, and 20.38 g (0.20 mol) of triethylamine to a 500 ml reaction flask. Heat to 75-80 °C and maintain for 5 hours, then cool to 20-25 °C within 2 hours. Adjust the pH to 2-3 with sulfuric acid. Filter and rinse with water until neutral. Dry at 60 °C for 10 hours to obtain 39.14 g of lentilana degradation impurities. The two-step yield was 76.0%. The HPLC purity was 96.1%, and the condensate content was 1.3%.

[0037] This invention is not limited to the above embodiments. Any simple or equivalent changes or modifications made to the above embodiments based on the technical essence of this invention shall fall within the scope of this invention.

Claims

1. A method for preparing lentilana to degrade impurities, characterized in that, Prepared via the following chemical reaction equation: ； Wherein, R1 = any one of methyl, ethyl, propyl, and isopropyl. The specific preparation steps for the above reaction equation are as follows: (1) Condensation: In a solvent, 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophen-2-carboxylic acid (abbreviated as S-isoxazolthiophenic acid), condensing agent, and glycine ester are added in a certain proportion. The mixture is heated and kept at the temperature. After the temperature is maintained, water is added to separate the layers. The organic layer is concentrated under reduced pressure to dryness to obtain the condensate. (2) Hydrolysis: The condensate is dissolved in a hydrolysis solvent, alkali is added in a certain proportion, the temperature is raised, the reaction is kept at the temperature, the temperature is lowered, acid is added to adjust the pH value to acidic, the mixture is filtered, washed with water to neutral, and dried to obtain lentilana degraded impurities.

2. The method for preparing lentilana-degraded impurities according to claim 1, characterized in that: In reaction step (1), the solvent is at least one selected from ethyl acetate, isopropyl acetate, butyl acetate, pentyl acetate, isoamyl acetate, and hexyl acetate, and the amount used is 2-15 times the weight of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisooxazol-3-yl]-thiophene-2-carboxylic acid; the condensing agent is at least one selected from dicyclohexylcarbodiimide, N,N-carbodiimidazole, diisopropylcarbodiimide, 1-hydroxybenzotriazole, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, and the amount used is 2-15 times the weight of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4, The molar ratio of 5-dihydroisoxazol-3-yl]-thiophen-2-carboxylic acid to the feed is 0.3-1.5:1; the glycine ester is at least one selected from glycine methyl ester, glycine ethyl ester, glycine propyl ester, and glycine isopropyl ester, and the molar ratio of its dosage to that of 3-methyl-5-[5-(3,4,5-trichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-thiophen-2-carboxylic acid is 1.05-2:1; the reaction temperature is 25-40℃; the reaction time is 3-7 hours; the amount of water added is 0.5-2 times the amount of solvent; the vacuum concentration temperature is 30-80℃, the vacuum degree is -0.06~-0.1MPa, and the endpoint of vacuum distillation is the absence of continuous distillate.

3. The method for preparing lentilana-degraded impurities according to claim 1, characterized in that: In reaction step (2), the hydrolysis solvent is at least one of methanol, ethanol, isopropanol, and acetone. The dosage is 2-10 times the weight of the condensate; the alkali is at least one of ammonia, triethylamine, potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, and lithium hydroxide, and the molar ratio of the dosage to the cyclized compound is 1.5-3:1; the heat preservation reaction temperature is 50-80℃; the heat preservation reaction time is 2-6 hours; the cooling endpoint temperature is 10-30℃, and the cooling time is 0.5-3 hours; the pH adjustment endpoint is 1-3, and the acid used is at least one of phosphoric acid, sulfuric acid, hydrochloric acid, nitric acid, phosphoric acid, formic acid, acetic acid, and propionic acid; the drying temperature is 50-80℃, and the drying time is 10-16 hours.

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