Method for preparing nitric oxide-donating prostaglandin analogues

Abstract

To provide a process for preparing the compound, hexanoic acid, 6-(nitrooxy)-, (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I).SOLUTION: The compound (I) can be efficiently prepared with high purity by coupling bimatoprost in a boronate-protected form with 6-(nitrooxy)hexanoyl chloride and removing the boronate-protecting group. A 6-(nitrooxy)hexanoyl chloride intermediate is prepared by ring-opening reaction of 2-caprolactone and subsequent nitration of a 6-hydroxyhexanoic acid potassium salt with a mixture of HNO3 and H2SO4 in dichloromethane.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an improved process for the large scale production of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester of formula (I). [Background technology]

[0002] Formula (I):

[0003] [ka] 6-(Nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester, represented by the formula (Impagnatiello F, Toris CB, Batugo M, Prasanna G, Borghi V, Bastia E, Ongini E, Krauss AHP; Invest Ophthalmol Vis Sci. 2015; 56:6558-64).

[0004] Methods for preparing compounds of formula (I) are disclosed in WO 2009 / 136281. WO 2009 / 136281 discloses a method for the synthesis of compound (I) specifically, and generally a method for the preparation of 15-alkyl nitrate esters of bimatoprost. WO 2009 / 136281 discloses a method for synthesizing the compound of formula (I) (Example B-1), in which a boronic acid protected form of bimatoprost (a compound of formula (II)) is reacted with 6-bromohexanoyl chloride to give a boronic acid protected form of bimatoprost 15-(6-bromohexanoyl) ester, which is converted to a nitric acid derivative with silver nitrate in acetonitrile, and deprotected / purified under reverse phase chromatography to produce the compound of formula (I).

[0005] [ka]

[0006] The main drawback of the above synthesis method is the use of more than equimolar amount of 6-bromohexanoyl chloride in the esterification reaction, which presents a potential mutagenic warning structure, and the use of silver nitrate in the last step, which generates a large amount of silver salt in wastewater.Another main drawback of this method is the formation of impurities and by-products, such as 15-(6-bromohexanoyl) ester of bimatoprost (compound (IX)) and 15-(6-chlorohexanoyl) ester of bimatoprost (compound (X)), which have similar chromatographic polarity, similar lipophilicity and / or solubility as compound (I), and are therefore difficult to remove even after multiple purifications.

[0007] [ka]

[0008] According to the procedure disclosed in WO 2009 / 136281, the 15-(6-bromohexanoyl) ester of bimatoprost (compound (IX)) is an impurity resulting from the incomplete reaction of compound (VIII) with silver nitrate after removal of the boronic acid protection. The 15-(6-chlorohexanoyl) ester of bimatoprost (compound (X)) is a by-product formed by the halogen exchange reaction of the bromine atom of the 15-(6-bromohexanoyl) ester of bimatoprost in boronic acid protected form (compound (VIII)) with the free chlorine anion of 4-dimethylaminopyridine hydrochloride formed during the esterification step. The 15-(6-chlorohexanoyl) ester of bimatoprost in boronic acid protected form (VIIIa) (Figure 3-Scheme 3) does not react with silver nitrate and produces compound (X) after removal of the protecting group.

[0009] WO 2009 / 136281 also discloses an alternative method for the preparation of 15-acylalkyl nitrate derivatives of bimatoprost (Examples N-1 and O-1). The synthesis involves reacting a boronic acid protected form of bimatoprost (compound of formula (II)) with an alkyl nitrate carboxylic acid chloride in the presence of resin-supported 4-dimethylaminopyridine (DMAP) (PS-DMAP), followed by removal of the boronic acid protecting group and purification using silica gel chromatography.

[0010] The above method avoids the use of 6-bromohexanoyl chloride and the removal of silver salts from the final product. However, this method presents another major drawback, the use of resin-supported 4-dimethylaminopyridine, which makes the method unsuitable and expensive for commercial scale-up. Furthermore, the nitrate alkyl carboxylic acid chloride is added in two successive steps and in high excess relative to the compound of formula (II), indeed the alkyl carboxylic acid chloride is added in an amount of about 2-4 equivalents.

[0011] WO 2009 / 136281 also discloses another method (Example Q1) for the preparation of 15-acyl alkyl nitrate derivatives of bimatoprost. In this method, the compounds are obtained by esterifying boronic acid protected form of bimatoprost (II) with excess alkyl nitrate-(p-nitrophenyl)-carboxylic acid in the presence of 4-dimethylaminopyridine.

[0012] The removal of unreacted alkyl nitrate-(p-nitrophenyl)-carboxylic acid using chromatographic methods and the removal of the by-product p-nitrophenol formed in equimolar amounts relative to the compound of formula (I) are the main drawbacks of this method.

[0013] WO 2016 / 155906 discloses 15-nitrooxy derivatives of fluprostenol and reports a method for the synthesis of 15-nitrooxy-hexyl ester of fluprostenol isopropyl ester, which was prepared by reacting boronic acid-protected form of fluprostenol isopropyl ester with (4-nitrophenyl)-6-nitrooxyhexanoate in the presence of an excess of 4-dimethylaminopyridine.

[0014] As reported above, the removal of unreacted alkyl nitrate-(p-nitrophenyl)-carboxylic acid, and especially the removal of the p-nitrophenol by-product, by chromatographic methods, is a major drawback of this method.

[0015] Over the past few years, various regulatory authorities have placed emphasis on the purity requirements and identification of impurities in Active Pharmaceutical Ingredients (API). Currently, impurities are considered as organic substances that can affect the efficacy and safety of the drug product other than the API. Therefore, the identification of each impurity and the quantification of the impurities, especially those with a mutagenicity alert structure, have become mandatory regulatory requirements. Furthermore, since the product is for pharmaceutical use, the range of industrially acceptable reagents, solvents, catalysts, etc. that can be used for the synthesis of the active ingredient is limited to those approved in the pharmaceutical industry.

[0016] The compound of formula (I) is an oil, which cannot be crystallized, making its purification difficult in large quantities, and therefore the presence of impurities is a major problem for large-scale production.The main sources of impurities are synthetic intermediates and by-products, so the purity of intermediates and the control of reaction conditions are important requirements for obtaining a final product with pharma- ceutically acceptable purity.

[0017] The prior art processes for preparing the compound of formula (I) have several shortcomings; for example, the use of bromohexanoyl chloride and reaction conditions result in the formation of the by-product 15-(6-chlorohexanoyl) ester of bimatoprost (compound (X)) which has a potential mutagenic warning structure; the use of silver nitrate for the preparation of intermediate alkyl nitrate chlorides or for the nitration of the boronic acid protected form 15-(6-bromohexanoyl) ester of bimatoprost (compound (VIII)) results in the management of large amounts of silver nitrate wastewater, and furthermore the metal content in the drug substance needs to meet certain acceptance standards.

[0018] There is therefore a need to provide the compound of formula (I) in high purity and good yield.

[0019] It has now been found that the compound of formula (I) can be produced in high purity by using a 6-(nitrooxy)hexanoyl chloride intermediate, which is efficiently prepared by a ring-opening reaction of caprolactone followed by nitration.

[0020] The present invention provides a large scale process for the production of the compound of formula (I) with high chemical purity, in particular with less than safe levels of the 15-(6-chlorohexanoyl) ester impurity of bimatoprost. [Brief description of the drawings]

[0021] [Figure 1] FIG. 1 shows steps 1 to 3 of a preferred method for producing compound (I). [Diagram 2] FIG. 1 shows steps 4 to 7 of a preferred method for producing compound (I). [Diagram 3] FIG. 1 shows the synthesis of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) according to the procedure disclosed in WO 2009 / 136281.

[0022] Description of the invention The object of the present invention is to provide a compound of formula (I):

[0023] [ka] A method for producing 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester represented by the following steps: a) Formula (II):

[0024] [ka] The compound represented by formula (IV):

[0025] [ka] in the presence of free 4-dimethylaminopyridine to obtain a compound of formula (III):

[0026] [ka] obtaining a compound represented by b) removing the boronic acid protecting group of the compound of formula (III) to obtain a compound of formula (I); The manufacturing method includes:

[0027] By free 4-dimethylaminopyridine (DMAP) is meant DMAP that is not bound to a resin.

[0028] Step a) is preferably carried out in an aprotic organic solvent, preferably selected from methyl tert-butyl ether, N,N-dimethylformamide or dichloromethane. Most preferably, the organic solvent is methyl tert-butyl ether.

[0029] The molar ratio of the compound of formula (II) to the 6-(nitrooxy)hexanoyl chloride of formula (IV) is preferably in the range of 1:1.4 to 1:1.6.

[0030] The molar ratio of the compound of formula (II) to 4-dimethylaminopyridine is preferably in the range of 1:2.0 to 1:2.4.

[0031] The reaction of step a) is carried out at a temperature ranging from 0° C. to room temperature.

[0032] The removal of the boronic acid protecting group (step b) is preferably carried out by reaction with methanol at a temperature between 17°C and 24°C.

[0033] 6-(nitrooxy)hexanoyl chloride of formula (IV) can be prepared by the following steps: i) Formula (V):

[0034] [ka] with an inorganic base selected from KOH, NaOH and LiOH to obtain a compound of formula (VI):

[0035] [ka] obtaining a 6-hydroxyhexanoate salt of the formula: ii) nitrating the compound of formula (VI) with a mixture of HNO3 and H2SO4 to give the compound of formula (VII):

[0036] [ka] obtaining 6-(nitrooxy)hexanoic acid represented by the formula: iii) converting the 6-(nitrooxy)hexanoic acid of formula (VII) with a chlorinating agent to 6-(nitrooxy)hexanoyl chloride of formula (IV) The composition is preferably obtained by a production method comprising the steps of:

[0037] The 6-(nitrooxy)hexanoyl chloride of formula (IV) obtained in step iii) may be reacted directly with a compound of formula (II) in step a) without further purification.

[0038] The inorganic base used in step i) is preferably potassium hydroxide.

[0039] Step i) is preferably carried out in a solvent selected from methanol, ethanol or isopropanol, most preferably methanol.

[0040] Steps ii) and iii) are carried out in dichloromethane.

[0041] The chlorinating reagent used in step iii) is oxalyl chloride.

[0042] The compound of formula (II) can be obtained by reacting bimatoprost with butylboronic acid.Preferably, the reaction is carried out in methyl tert-butyl ether as a solvent.

[0043] A preferred process for preparing compound (I) is illustrated in more detail in Schemes 1 and 2 (FIGS. 1 and 2), which comprises the following steps: Step 1) reacting 2-caprolactone of formula (V) with potassium hydroxide in methanol to obtain potassium 6-hydroxyhexanoate (a compound of formula (VI) where M is potassium); Step 2) reacting 6-hydroxyhexanoic acid potassium salt with a mixture of HNO3 and H2SO4 in dichloromethane to obtain 6-(nitrooxy)hexanoic acid (compound of formula (VII)); step 3) reacting 6-(nitrooxy)hexanoic acid with oxalyl chloride to obtain 6-(nitrooxy)hexanoyl chloride (compound of formula (IV)), which is used without further purification; step 4) reacting bimatoprost with butylboronic acid (1.1-1.8 equivalents) in methyl tert-butyl ether (MTBE) at a temperature of about 40° C., followed by removing water by azeotropic distillation to obtain bimatoprost boronate (compound of formula (II)); step 5) reacting bimatoprost borate (compound of formula (II)) with 6-(nitrooxy)hexanoyl chloride (IV) (1.4-1.6 equivalents) in methyl tert-butyl ether in the presence of 4-dimethylaminopyridine (2.0-2.4 equivalents) at a temperature ranging from 0° C. to about room temperature to obtain compound of formula (III); Step 6) reacting the compound of formula (III) with methanol at room temperature for removing the protecting group to obtain a crude compound of formula (I); Step 7) Purifying the crude compound of formula (I) to obtain compound (I) having a chemical purity of greater than 99%. Includes.

[0044] The process of the present invention is characterized in that the intermediate 6-(nitrooxy)hexanoyl chloride (compound (IV)) is prepared in high chemical purity and high yield by ring-opening reaction of 2-caprolactone.

[0045] The experimental procedures for the steps of the manufacturing method of the present invention are detailed below. All steps are carried out under a nitrogen atmosphere.

[0046] 6-(Nitrooxy)hexanoyl chloride (IV) is prepared in high purity and yield starting from 2-caprolactone (FIG. 1-Scheme 1); the synthesis includes the following steps 1) to 7): Step 1) A solution of potassium hydroxide (1 equivalent) in methanol is added dropwise to a solution of 2-caprolactone (1 equivalent) in methanol; the mixture is cooled to about 5°C-20°C and stirred at 15°C-20°C for about 5 hours after the addition is complete; the solvent is removed (temperature is below 40°C), the crude product is slurried in methyl tert-butyl ether, and 6-hydroxyhexanoic acid potassium salt (VI) is filtered, washed with methyl tert-butyl ether and dried. 6-hydroxyhexanoic acid potassium salt (VI) is obtained in 95% yield and 98.5% purity ( 1 H-NMR and HCl assay); Step 2) 6-hydroxyhexanoic acid potassium salt (VI) (1 eq.) is added in portions to a mixture of HNO3 (4.6 eq.) and H2SO4 (3.1 eq.) in dichloromethane cooled at 0°C-5°C under nitrogen over a period of about 30 minutes while maintaining the temperature below 10°C; 1 The resulting mixture is stirred at 0°C-10°C for about 2-3 hours while monitoring the completion of the reaction by H-NMR analysis; the mixture is cooled to a temperature of 0°C-5°C and saturated aqueous sodium chloride solution is added dropwise in about 20 minutes; the reaction mixture is maintained at a temperature below 10°C; the organic layer is dried over anhydrous sodium sulfate and the solvent is removed to give 6-(nitrooxy)hexanoic acid (VII) in 86-88% yield and 97% HPLC purity; Step 3) N,N-dimethylformamide and oxalyl chloride are added dropwise to a solution of 6-(nitrooxy)hexanoic acid (VII) in dichloromethane over 1 hour while maintaining the temperature of the solution at 0°C-5°C, then the mixture is stirred at 15°C-30°C for 24 hours; the solvent is evaporated to give 6-(nitrooxy)hexanoyl chloride (IV) in 88-97% yield, which is used without further purification.

[0047] The esterification method between bimatoprost and 6-(nitrooxy)hexanoyl chloride includes the synthetic steps disclosed below: Step 4) Bimatoprost is added to methyl tert-butyl ether and the resulting solution is cooled to about 15°C to 18°C; n-butylboronic acid (1.11 to 1.18 equivalents) is then added in one portion and the mixture is stirred at 40°C for about 1 to 1.5 hours. 1 H NMR analysis; the reaction mixture is cooled to about 20° C.-25° C., filtered and the water formed is removed by azeotropic distillation of methyl tert-butyl ether at a temperature not exceeding 40° C. until the water content is not more than 0.25%, to give crude bimatoprost boronate (II), which is used directly in the next step; Step 5) Crude bimatoprost borate is added to methyl tert-butyl ether, the resulting solution is cooled to about 0°C to 5°C, 4-dimethylaminopyridine (about 2.1 to 2.3 equivalents) is added, and 6-(nitrooxy)hexanoyl chloride (compound (IV)) (1.5 equivalents) dissolved in methyl tert-butyl ether is added dropwise while maintaining the temperature of the mixture at about 0°C to 5°C. After the addition, the mixture is stirred at about 0° C.-5° C. for up to 4 hours and then up to 15° C.-20° C. overnight; completion of the reaction is monitored by HPLC analysis; (1S,2E)-3-{(6R,7R)-3-butyl-7-[(2Z)-7-(ethylamino)-7-oxohept-2-en-1-yl]-2,4-dioxa-3-borabicyclo[3.2.1]oct-6-yl}-1-(2-phenylethyl)-prop-2-en-1-yl 6-(nitrooxy)hexanoate (compound (III)) is isolated by standard work-up methods (one example of the work-up method is described in Example 1); Step 6) Compound (III) is dissolved in methanol, and the resulting solution is stirred at 17°C to 25°C for about 18 hours; the conversion of compound (III) to compound (I) is 1 The reaction is monitored by H NMR. When the reaction has stopped, the mixture is evaporated and redissolved in fresh methanol until complete conversion; the reaction mixture is then concentrated under vacuum at a temperature below 40° C. and the crude compound (I) is isolated by standard work-up methods; Step 7) The obtained crude product (I) is purified by column chromatography using a silica gel column and a mixed solvent of dichloromethane and methanol to give compound (I) with an overall yield of more than 60% from bimatoprost and a purity of more than 99%.

[0048] The process of the present invention provides the compound of formula (I) in high yield and purity while reducing the amount of by-products, in particular (S,E)-1-((1R,2R,3S,5R)-2-((Z)-7-(ethylamino)-7-oxohept-2-enyl)-3,5-dihydroxycyclopentyl)-5-phenylpent-1-en-3-yl 6-chlorohexanoate (compound (X)), which has a potential mutagenic warning structure.

[0049] The above advantages make the manufacturing method of the present invention a cost-effective manufacturing method that can be easily transferred to an industrial scale. EXAMPLES

[0050] All synthetic steps described below were carried out under a nitrogen atmosphere. Example 1

[0051] Synthesis of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) (batch 1) Synthesis of 6-(nitrooxy)hexanoyl chloride(IV) Step 1: Synthesis of 6-hydroxyhexanoic acid potassium salt (compound (VI)) A solution of potassium hydroxide (131.9 g, 0.98 eq.) in methanol (1250 ml, 5 volumes) was prepared under cooling at 15° C.-20° C. 250 g of 2-caprolactone (1 eq.) and methanol (625 ml, 2.5 volumes) were introduced into a 3 L three-necked round bottom flask. The mixture was stirred until dissolved. Methanolic potassium hydroxide solution was added within 0.5 hours at 5° C.-20° C. The mixture was stirred at 15° C.-20° C. for 4.5 hours. The reaction mixture was concentrated under vacuum (at a temperature below 40° C.) to give crude 6-hydroxyhexanoic acid potassium salt (489.55 g). The crude product was reslurried in methyl tert-butyl ether (1250 ml, 5 vol) at 20°C-25°C for 4 hours, filtered through a Pore 3 filter, washed with methyl tert-butyl ether (2 x 250 ml, 2 x 1 vol) and dried under vacuum (temperature below 40°C) to give 6-hydroxyhexanoic acid potassium salt (353.04 g) in 95.7% yield. Melting point 208°C. The product was 1 Analyzed by H-NMR and HCl assay. Step 2: Synthesis of 6-(nitrooxy)hexane (VII) The reaction is carried out on a 100 g scale to control the reaction temperature and shorten the addition time of the nitration mixture. The reaction is repeated to obtain the required amount of 6-(nitrooxy)hexanoic acid. Fuming HNO3 (4.6 equiv.) was added to concentrated H2SO4 (3.1 equiv.) at a temperature between 0°C and 5°C in 14 min, followed by CH2Cl2 (20 vol.) at a temperature between 0°C and 5°C in 12 min. 6-Hydroxyhexanoic acid potassium salt (101.29 g, 1 equiv.) was added in portions at a temperature below 10°C in 28 min. The mixture was stirred at 0°C to 10°C for 2.2 h to complete the reaction. 1 Monitoring by H-NMR showed 99.9% conversion. The mixture was cooled to 0-5°C and saturated aqueous sodium chloride solution (286.71 g in 910 mL, 10 volumes) was carefully added within 17 min at a temperature below 10°C. After filtration of the insolubles, the organic layer was decanted, dried over sodium sulfate and concentrated under vacuum (at a temperature below 40°C) to give 6-(nitrooxy)hexanoic acid in 87.7% yield and 97.0% HPLC purity. Step 3: Synthesis of 6-(nitrooxy)hexanoyl chloride (IV) 6-(Nitrooxy)hexanoic acid (230 g, 1 equiv.) was dissolved in dichloromethane (150 mL, 5 vol.). The resulting solution was filtered through glass fiber, washed with dichloromethane (1×50 mL, 0.65 vol.) and analyzed by Karl Fischer method (water content=0.016%). The filtrate was cooled to 0° C.-5° C. under nitrogen. N,N-dimethylformamide (1.35 mL, 0.0059 vol.) and oxalyl chloride (108.5 mL, 1 equiv.) were then added at 0° C.-5° C. within 34 min. The reaction mixture was stirred at 0° C.-5° C. for 3.5 h and at 15° C.-20° C. for 14 h. TLC monitoring revealed the completion of the reaction. The medium was concentrated under vacuum (temperature below 40° C.) and coevaporated with dichloromethane (4×1 L, 4×4.35 vol) to give 6-(nitrooxy)hexanoyl chloride (240.0 lg) in 97.7% yield. Step 4: Preparation of (Z)-7-[(1S,5R,6R,7R)-3-butyl-6-[(E,3S)-3-hydroxy-5-phenyl-pent-1-enyl]-2,4-dioxa-3-borabicyclo[3.2.1]octan-7-yl]-N-ethyl-hept-5-enamide (II) The flask was charged with methyl tert-butyl ether (2800 mL, 14 vol). Bimatoprost (200 g, 1 eq) was added and the apparatus was rinsed with methyl tert-butyl ether (200 mL, 1 vol). Butyl boronic acid (58.94 g, 1.13 eq) was added in one portion to the resulting suspension and the apparatus was rinsed with methyl tert-butyl ether (200 mL, 1 vol). The mixture was heated to 40° C. for 1 h. Conversion to >97% 1 The reaction was monitored by 1 H NMR. The reaction mixture was cooled to 20°C-25°C, clarified through a glass filter and washed with methyl tert-butyl ether (200 mL, 1 volume). The filtrate was charged into a 4 L three-necked round bottom flask, the apparatus was rinsed with methyl tert-butyl ether (100 mL, 0.5 volume) and the medium was heated under vacuum at a temperature of about 40°C for azeotropic distillation. Rinsing with methyl tert-butyl ether and azeotropic distillation were continued until the water content of (Z)-7-[(1S,5R,6R,7R)-3-butyl-6-[(E,3S)-3-hydroxy-5-phenyl-pent-1-enyl]-2,4-dioxa-3-borabicyclo[3.2.1]octan-7-yl]-N-ethyl-hept-5-enamide (compound (II)) was 0.25% or less. The compound of formula (II) was obtained in quantitative yield (281.22 g). Step 5: Preparation of (1S,2E)-3-{(6R,7R)-3-butyl-7-[(2Z)-7-(ethylamino)-7-oxohept-2-en-1-yl]-2,4-dioxa-3-borabicyclo[3.2.1]oct-6-yl}-1-(2-phenylethyl)-prop-2-en-1-yl 6-(nitrooxy)hexanoate (III) Methyl tert-butyl ether (3196 mL, 11.6 vol) was charged to a 4 L three-necked round bottom flask under nitrogen. (Z)-7-[(1S,5R,6R,7R)-3-butyl-6-[(E,3S)-3-hydroxy-5-phenyl-pent-1-enyl]-2,4-dioxa-3-borabicyclo[3.2.1]octan-7-yl]-N-ethyl-hept-5-enamide (compound (II)) (276.48 g crude, 1 eq) was added and the apparatus was rinsed with methyl tert-butyl ether (398 mL, 1.44 vol). The resulting solution was analyzed by Karl Fischer (water content = 0.072%) and cooled to 0°C-5°C. 4-Dimethylaminopyridine (138.5 g, 2.27 eq) was added in one portion. A solution of 6-(nitrooxy)hexanoyl chloride (172.6 g, 1.5 eq) in methyl tert-butyl ether (508 mL, 1.84 vol) was added dropwise within 1 h at 0°C-5°C. The addition funnel was rinsed with methyl tert-butyl ether (32 mL, 0.12 vol). After stirring for 24 min at 0°C-5°C, HPLC monitoring revealed 97.8% conversion. The mixture was stirred at 15-20°C for 14.5 h. HPLC monitoring revealed 99.8% conversion. The reaction mixture was cooled at 0°C-5°C and deionized water (1351 mL, 4.89 vol) was added within 20 min at a maximum temperature of 15°C. The mixture was stirred for 5 minutes and decanted. The aqueous layer was analyzed and then discarded. An aqueous solution of 1 N hydrochloric acid was prepared by mixing deionized water (493 mL, 1.78 vol) and 11.6 N hydrochloric acid (46.6 mL, 1.08 eq). The organic layer was washed with the hydrochloric acid solution. The aqueous layer was analyzed and then discarded. The organic layer was washed first with deionized water (1351 mL, 4.88 vol) and then with saturated sodium chloride solution (3×1177 mL, 3×4.25 vol) previously prepared by mixing deionized water (3530 mL, 12.8 vol) and sodium chloride (1175 g, 425% w / w). The aqueous layer (pH=4 after the last wash) was analyzed and discarded. The organic layer was dried over sodium sulfate (240 g, 86.8% w / w), washed with methyl tert-butyl ether (481 mL, 1.74 vol) and concentrated under vacuum to give (1S,2E)-3-{(6R,7R)-3-butyl-7-[(2Z)-7-(ethylamino)-7-oxohept-2-en-1-yl]-2,4-dioxa-3-borabicyclo[3.2.1]oct-6-yl}-1-(2-phenylethyl)-prop-2-en-1-yl 6-(nitrooxy)hexanoate (compound (III)) in quantitative yield (327 g). Step 6: Synthesis of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) (crude compound). (1S,2E)-3-{(6R,7R)-3-butyl-7-[(2Z)-7-(ethylamino)-7-oxohept-2-en-1-yl]-2,4-dioxa-3-borabicyclo[3.2.1]oct-6-yl}-1-(2-phenylethyl)-prop-2-en-1-yl 6-(nitrooxy)hexanoate (compound (III)) (325.4 g crude, 1 eq.) was dissolved in methanol (2873 mL, 8.83 vol.). The resulting solution was charged to a flask and the apparatus was rinsed with methanol (918 mL, 2.82 vol.). The mixture was added to 100 ml of methanol (100 ml). The resulting ... 1 The reaction was monitored by H-NMR while stirring for 2.40 hours at 17° C.-24° C. The mixture was stirred at 16° C.-20° C. for an additional 15 hours. 1 As no progress was observed by H-NMR monitoring, the methanol was removed under vacuum at 35° C.-40° C. To the residue was added methanol (918 mL, 2.82 vol) and the mixture was stirred at 20° C.-25° C. for 4 h. 1H-NMR monitoring showed 94.6% conversion. The reaction mixture was concentrated under vacuum at a temperature below 40° C. The residue was dissolved in methyl tert-butyl ether (3530 mL, 10.85 vol). The resulting solution was washed with deionized water (1770 mL, 5.44 vol). The aqueous layer (pH=7) was discarded. A solution of sodium chloride (1045 g, 321% w / w) in deionized water (3134 mL, 9.63 vol) was prepared. The organic layer was washed with sodium chloride solution (2×1567 mL, 2×4.82 vol). The aqueous layer was discarded. The organic layer was dried over sodium sulfate (320 g, 98% w / w), washed with methyl tert-butyl ether (640 mL, 1.97 vol) and concentrated under vacuum at a temperature below 40° C. to give crude compound (I) (292.77 g) in quantitative yield. Step 7: Purification of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I). The mixture (crude compound (I)) obtained in step 6) was divided into four portions and purified on Combiflash using silica gel column (750 g×4, 10.34 vol) and dichloromethane / methanol as eluent with a gradient of 100:0 to 95:5. The fractions were monitored by TLC and analyzed by HPLC (area %). The fractions with HPLC (area %) ≧98% were mixed and concentrated under vacuum below 50° C. to give 188.83 g of compound (I), which was further charged into a 4 L three-necked round bottom flask, dissolved in absolute ethanol (1510 mL, 8 vol), and the apparatus was rinsed with absolute ethanol (378 mL, 2 vol). Activated charcoal (19 g, 10% w / w) was added and the mixture was stirred at 20° C. to 25° C. for 0.5 h. The activated charcoal was filtered and washed with absolute ethanol (189 mL, 1 vol). The filtrate was concentrated under industrial vacuum at 45°C-50°C for 2 h, then concentrated under high vacuum at 45°C-50°C for 5 h. 1 Monitored by 1 H NMR showed no residual solvent. 6-(Nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) (174.4 g) was obtained in an overall yield of 63% from compound (II). The HPLC purity was 99.47%. Two other batches (2 and 3) of compound of formula (I) were prepared by the same procedure as described in Example 1, and the purity of compound of formula (I) and the total yield from compound (II) of the three batches are reported in Table 1 below.

[0052] [Table 1] Example 2

[0053] (Comparative Example) Synthesis of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) according to the procedure disclosed in WO 2009 / 136281 (Figure 3-Scheme 3). Step A: Preparation of (Z)-7-[(1S,5R,6R,7R)-3-butyl-6-[(E,3S)-3-hydroxy-5-phenyl-pent-1-enyl]-2,4-dioxa-3-borabicyclo[3.2.1]octan-7-yl]-N-ethyl-hept-5-enamide (II) Butylboronic acid (1.129 equiv.) was added to a solution of bimatoprost (1 g, 1 equiv.) in dichloromethane (16 vol.). 1The mixture was heated to 40° C. for 1 h while monitoring the progress of the reaction by H-NMR. The solvent was removed under reduced pressure for 2 h. Dichloromethane (16 vol) was added and the mixture was heated to 40° C. for a further 1 h. The solvent was removed under pressure for 40 min. Dichloromethane (16 vol) was added and the mixture was heated to 40° C. for 16 h. The solvent was evaporated and the crude product was dried under high vacuum at 40° C. for 3 h to yield compound (II) in quantitative yield, which was used in the next step without further purification. MS: m / z=438[M+H]+ Step B: Synthesis of (S,E)-1-((1S,5R,6R,7S)-3-butyl-7-((Z)-7-(ethylamino)-7-oxohept-2-en-1-yl)-2,4-dioxa-3-borabicyclo[3.2.1]octan-6-yl)-5-phenylpent-1-en-3-yl 6-bromohexanoate (VIII). 4-Dimethylaminopyridine (1.1 eq) and 6-bromohexanoyl chloride (1.15 eq) were added to a solution of compound (II) (0.8 g, 1 eq) in dichloromethane (15.3 vol) cooled to 0-5° C. The mixture was stirred at 0-5° C. for 0.5 h and at 20-25° C. for 16 h. 4-Dimethylaminopyridine (0.25 eq.) and 6-bromohexanoyl chloride (0.25 eq.) were added and the mixture was stirred for a further 19 h. The reaction was allowed to proceed until the conversion was complete. 1 The mixture was diluted with dichloromethane (15.3 vol) and the organic solution was washed with deionized water (6.25 vol) and brine (6.25 vol). The organic phase was dried over Na2SO4 and concentrated in vacuo to give compound (VIII) as a pale yellow oil, which was used in the next step without further purification (calculated as quantitative yield). Step C: Synthesis of 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester (I) Silver nitrate (3.72 equiv.) was added to a solution of compound (VIII) (0.8 g, 1 equiv.) in acetonitrile (9.4 vol.). The mixture was stirred at 20° C.-25° C. for 18 h. The conversion was confirmed by the addition of 1.0% ethyl acetate in DMSO. 1 H-NMR was used to monitor. Silver nitrate (0.5 eq) was added and the mixture was stirred for an additional 20 h, at which time HPLC showed 99.7% conversion. The mixture was filtered through a Whatman filter. The filtrate was concentrated under vacuum. The residue was dissolved in ethyl acetate (30 volumes). The organic phase was washed with deionized water (5 volumes) and brine (5 volumes). After drying over Na2SO4, the layer was concentrated under vacuum. The residue was chromatographed on a silica gel column using dichloromethane / methanol (95:5) as eluent. The fractions were monitored by TLC, and only the fractions showing a single point were combined and concentrated under vacuum at a temperature below 40°C to produce compound (I) in 86% overall yield with 4.27% bimatoprost. Reverse phase HPLC (area %) revealed that the purity of compound (I) was 77% and the content of 15-(6-chloridohexanoyl) ester of bimatoprost (compound (X)) was 8.34%.

[0054] [Table 2]

[0055] Table 2 reports the yield of compound (I) and the main impurities formed during its preparation by the process according to the invention (Example 1) and by the process disclosed in WO 2009 / 136281 (Example 2). Compound (X) is the 15-(6-chlorohexanoyl) ester of bimatoprost, which has a structural alert for potential mutagenicity. The results show that the preparation method of the present invention provides compound (I) having a chemical purity of more than 99% and a compound (X) content of 0.15% to 0.26%, while the preparation method disclosed in the prior art results in compound (I) having a chemical purity of 77% and a compound (X) content of 8.34% (i.e., more than 30 times the amount of compound (X) formed by the preparation method of the present invention). The results demonstrate that the manufacturing method of the present invention represents an improvement over the manufacturing methods described in the prior art.

Claims

[Claim 1] 1. A 6-(nitrooxy)-hexanoic acid (1S,2E)-3-[(1R,2R,3S,5R)-2-[(2Z)-7-(ethylamino)-7-oxo-2-hepten-1-yl]-3,5-dihydroxycyclopentyl]-1-(2-phenylethyl)-2-propen-1-yl ester composition having a chemical purity of greater than 99% as determined by HPLC analysis and comprising (S,E)-1-((1R,2R,3S,5R)-2-((Z)-7-(ethylamino)-7-oxohept-2-enyl)-3,5-dihydroxycyclopentyl)-5-phenylpent-1-en-3-yl 6-chlorohexanoate in an amount between 0.15% and 0.26%.

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