Method for synthesizing cationic lipids

Abstract

The present application provides methods for synthesizing cationic lipids of Formula I that can be used to synthesize lipid-soluble compositions for targeting and enhancing the activity of therapeutic molecules, including siRNA.

Description

Technical Field

[0001] This application provides a method for synthesizing cationic lipids, which can be used to synthesize lipid-soluble compositions for targeting and enhancing the activity of therapeutic molecules, including siRNA. Background Technology

[0002] Many techniques can be used to deliver therapeutic agents such as siRNA into cells, including the use of viral and non-viral transfection systems. Non-viral transfection systems can include, for example, polymers, lipids, liposomes, micelles, dendritic polymers, and nanomaterials. Examples of polymers previously studied for cell transfection include cationic polymers such as poly(L-lysine) (PLL), polyethyleneimine (PEI), chitosan, and poly(2-dimethylamino)ethyl methacrylate (pDMAEMA). Each type of system has its own advantages and disadvantages. For example, viral systems can produce high transfection efficiency but may be less safe than some non-viral systems. Additionally, the preparation of viral systems can be complex and / or expensive. Non-viral transfection systems, such as cationic polymers, have been reported to transfer plasmid DNA into cells. However, some disadvantages of using cationic polymers include their cytotoxicity and / or their lack of stability. To date, the amino-alkylhydroxyl (N-alkyl-OH) moiety of compounds of Formula I has been found to impart properties to formulations not previously seen in other previously reported cationic lipids. Formulations containing the cationic lipid of formula I resulted in a superior reduction in protein expression compared to formulations that did not contain formula I. Particularly surprising was the ability of formulations containing the cationic lipid of formula I to reduce HSP47 expression.

[0003] One such example of a cationic lipid is 2-(bis(2-(tetradecanoyloxy)ethyl)amino)-N-(2-hydroxyethyl)-N,N-dimethyl-2-oxoethylamineonium bromide (HEDC), as disclosed in U.S. Patent No. 9,242,001. Previous synthetic strategies for producing HEDC and similar compounds have been plagued by low yields, inconvenient post-processing procedures, and large amounts of byproducts.

[0004] There is still a need for synthetic methods for producing these cationic lipids that promote increased product yields, low byproduct yields, and use more readily available synthetic techniques. Summary of the Invention

[0005] In one aspect, this application provides a method for producing HEDC.

[0006] In one embodiment, this application provides a method for synthesizing compounds of formula I.

[0007]

[0008] Where a is an integer between 8 and 14;

[0009] b is an integer between 1 and 3; and

[0010] Z is a counter ion.

[0011] The method includes

[0012] a) Make the compound of formula II

[0013]

[0014] Where R is a protecting group;

[0015] Compounds of Formula III

[0016]

[0017] Where X is a halogen.

[0018] The reaction forms a compound of formula IV.

[0019]

[0020] b) Mixing the compound of formula IV with the compound of formula V

[0021]

[0022] The reaction forms a compound of formula VI.

[0023]

[0024] as well as

[0025] c) Reacting the compound of formula VI with bromoethanol under coupling conditions to form the compound of formula I.

[0026] In yet another implementation, a is 12.

[0027] In yet another implementation, b is 2.

[0028] In another embodiment, R is independently selected from carboxybenzyl, p-methoxybenzylcarbonyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, acetyl, trifluoroacetyl, benzoyl, benzyl, carbamate, p-methoxybenzyl, 3,4-dimethoxybenzyl, p-methoxybenzyl, toluenesulfonyl, trichloroethyl chloroformate, (4-nitrophenyl)sulfonyl, methyl, ethyl, propyl, n-butyl, tert-butyl, succinimide, 2,6-dimethylphenol, 2,6-diisopropylphenol, 2,6-ditert-butylphenol, trimethylsilyl, allyl, 1,1-dimethylallyl, 2,2,2-trifluoroethyl, phenyl, and 4-methoxybenzyl.

[0029] In yet another embodiment, R is a tert-butoxycarbonyl group.

[0030] In yet another embodiment, X is Cl or Br. In yet another embodiment, X is Cl.

[0031] In yet another implementation scheme, Z is Cl - or Br - In another implementation, Z is Br - .

[0032] In yet another embodiment, the yield of step a) is at least about 75%. In yet another embodiment, the yield of step b) is at least about 75%.

[0033] In yet another embodiment, in step c), bromoethanol is used in amounts between about 2 equivalents and about 4 equivalents, and the compound of formula VI is used in an amount of about 1 equivalent. In yet another embodiment, in step c), bromoethanol is used in an amount of about 2.2 equivalents, and the compound of formula VI is used in an amount of about 1 equivalent.

[0034] In yet another embodiment, the compound of formula IV is isolated in solid form. In yet another embodiment, the compound of formula VI is isolated in crystalline solid form. In yet another embodiment, the compound of formula VI is isolated in crystalline solid form with a yield of 80%-90%.

[0035] In yet another embodiment, the compound of formula VI is isolated in solid form. In yet another embodiment, the compound of formula VI is isolated in crystalline solid form. In yet another embodiment, the compound of formula VI is isolated in crystalline solid form with a yield of 80%-95%.

[0036] In yet another embodiment, the compound of formula I is

[0037]

[0038] In yet another embodiment, the compound of formula I is produced by the compound of formula II in a yield of at least about 60%.

[0039] In yet another embodiment, the compound of formula I is produced by the compound of formula II in a yield of at least about 65%.

[0040] In another embodiment, the compound of formula I is prepared by steps a)-c). Detailed Implementation

[0041] Throughout the specification and appended claims, the given chemical formula or name shall include all its stereo and optical isomers and racemates, where such isomers are present. Unless otherwise indicated, all chiral (enantiomers and diastereomers) and racemic forms are within the scope of this invention. Many geometric isomers of C=C double bonds, C=N double bonds, cyclic systems, etc., may also be present in the compounds, and all such stable isomers are contemplated in this invention. Cis- and trans- (or E- and Z-) geometric isomers of the compounds of this invention are described and can be separated into mixtures of isomers or isolated isomeric forms. The compounds of this invention can be separated in optically active or racemic forms. Optically active forms can be prepared by resolving racemic forms or by synthesis from optically active starting materials. All methods used to prepare the compounds of this invention and the intermediates prepared therein are considered part of this invention. When preparing enantiomers or diastereomers, they can be separated by conventional methods (e.g., by chromatography or fractional crystallization).

[0042] Depending on the method conditions, the end products of this invention can be obtained in free (neutral) or salt form. Both the free form and the salt of these end products are within the scope of this invention. If desired, one form of the compound can be converted to another. A free base or acid can be converted to a salt; a salt can be converted to a free compound or another salt; a mixture of isomers of the compounds of this invention can be separated into individual isomers. The compounds of this invention, their free forms, and salts can exist in a variety of tautomeric forms, wherein hydrogen atoms are transposed to other parts of the molecule, and thus the chemical bonds between the atoms of the molecule are rearranged. It should be understood that all tautomeric forms, wherever they may exist, are included within the scope of this invention.

[0043] The term "stereoisomer" refers to isomers that have the same composition but differ in the spatial arrangement of their atoms. Enantiomers and diastereomers are examples of stereoisomers. The term "enantiomer" refers to one of a pair of molecular species that are mirror images of each other and cannot be superimposed. The term "diastereomer" refers to a stereoisomer that is not a mirror image. The term "racemic mixture" or "racemic mixture" refers to a composition consisting of two enantiomer species in equimolar amounts, wherein the composition is not optically active. It is well understood in the art that the stereochemistry of the product can be controlled by selecting the stereochemistry of the starting material, and that the stereochemistry of the product can be altered by changing the stereochemistry of the starting material. It is also well understood in the art how to separate racemic mixtures such that the stereochemical purity of the product is >99%.

[0044] The abbreviations used in this article are defined as follows: "°C" represents degrees Celsius, "eq" represents equivalent, "g" represents gram, "mg" represents milligram, "L" represents liter, "mL" represents milliliter, "μL" represents microliter, "N" represents equivalent concentration, "M" represents molar concentration, "mmol" represents millimole, "min" represents minute, "h" represents hour, "rt" represents room temperature, "RT" represents retention time, "conc." represents concentrate, "sat" or "saturated" represents saturation, "MW" represents molecular weight, "ee" represents enantiomer excess, "MS" or "Mass Spec" represents mass spectrometry, "ESI" represents electrospray ionization mass spectrometry, "HR" represents high resolution, "HRMS" represents high resolution mass spectrometry, "LCMS" represents liquid chromatography-mass spectrometry, "HPLC" represents high performance liquid chromatography, and "NMR" represents nuclear magnetic resonance spectroscopy. 1 "H" represents the proton, and "D", "L", "α", "β", "R", "S", "E" and "Z" are stereochemical names familiar to those skilled in the art.

[0045] This document provides a method for synthesizing cationic lipids. In one embodiment, the method is a method for synthesizing compounds of formula I. In one embodiment, the method is a method for synthesizing compounds of formula I starting from compounds of formula II.

[0046] Step a)

[0047] In one implementation, step a) includes

[0048] Compounds of Formula II

[0049]

[0050] Where R is a protecting group;

[0051] Compounds of Formula III

[0052]

[0053] Where a is an integer between 8 and 14; and

[0054] X is a halogen.

[0055] The reaction forms a compound of formula IV.

[0056]

[0057] In one embodiment, R is an amine protecting group. In another embodiment, R is selected from carboxybenzyl, p-methoxybenzylcarbonyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, acetyl, trifluoroacetyl, benzoyl, benzyl, carbamate, p-methoxybenzyl, 3,4-dimethoxybenzyl, p-methoxybenzyl, toluenesulfonyl, trichloroethyl chloroformate, and (4-nitrophenyl)sulfonyl. In one embodiment, R is tert-butoxycarbonyl.

[0058] In one embodiment, a is an integer from 8 to 14. In another embodiment, a is an integer from 8 to 9, 8 to 10, 8 to 11, 8 to 12, or 8 to 13. In one embodiment, a is an integer from 1 to 6. In another embodiment, a is an integer from 2 to 10, 3 to 10, 4 to 10, 5 to 10, 6 to 10, 7 to 10, 8 to 10, or 9 to 10. In another embodiment, a is an integer from 9 to 13 or 10 to 12. In one embodiment, a is 8. In one embodiment, a is 9. In one embodiment, a is 10. In one embodiment, a is 11. In one embodiment, a is 12. In one embodiment, a is 13. In one embodiment, a is 14.

[0059] In one embodiment, X is a halogen. In another embodiment, X is selected from fluorine (F), chlorine (Cl), bromine (Br), iodine (I), astatine (At), and tennessine (Ts). In yet another embodiment, X is F. In yet another embodiment, X is Cl. In yet another embodiment, X is Br. In yet another embodiment, X is I.

[0060] In one embodiment, step a) is carried out in a solvent. In one embodiment, step a) is carried out in a solvent mixture. In one embodiment, at least one solvent is nonpolar. In one embodiment, the one or more solvents are selected from pentane, hexane, cyclohexane, benzene, toluene, heptane, MTBE, cyclopropyl methyl ether, and diethyl ether. In one embodiment, the solvent is not a di or trihalogenated solvent. In one embodiment, step b) is carried out in toluene.

[0061] In one embodiment, the compound of formula IV is separated in crystalline solid form. In one embodiment, step a) does not require chromatography for separation or purification.

[0062] In one embodiment, the compound of formula IV is separated in crystalline solid form. In one embodiment, step a) does not require chromatography for separation or purification. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula IV, based on liquid chromatographic area percentage (LCAP), is between about 95% and 99.9%. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula IV, based on LCAP, is at least about 95%.

[0063] In one embodiment, the crystalline compound of formula IV is produced from step a) in a yield between about 70% and 95%. In another embodiment, the compound of formula IV is produced in a yield between about 70% and 85%, or between about 70% and 75%. In yet another embodiment, the compound of formula IV is produced in a yield between about 80% and about 95%, or between about 90% and 95%. In yet another embodiment, the compound of formula IV is produced in a yield of about 70%, about 75%, about 80%, about 85%, about 90%, or about 95%. In yet another embodiment, the compound of formula IV is produced in a yield of about 80%. In one embodiment, the compound of formula IV is produced from step a) in a yield of at least about 65%, at least 70%, at least 75%, at least about 80%, or at least about 85%. In another embodiment, formula IV is produced from step a) in a yield of at least about 80%.

[0064] In one embodiment, step a) is a simplified two-step process. The first reaction is the bismyristylation of the compound of formula I catalyzed by N-methylmorpholine. After the reaction is complete, the mixture is washed with an aqueous solution of 1N acetic acid to remove unreacted starting material and salt, and then subjected to dry distillation. The second reaction is the N-Boc-deprotection reaction followed by salt formation with methanesulfonic acid (MSA).

[0065] Step b

[0066] In one embodiment, step b includes reacting the compound of formula IV with the compound of formula V.

[0067]

[0068] Where b is an integer from 1 to 3;

[0069] The reaction forms a compound of formula VI.

[0070]

[0071] Where a is an integer from 8 to 14; and

[0072] b is an integer between 1 and 3.

[0073] In one implementation, a and b are as described with respect to step a). In one implementation, a is 12 and b is 2.

[0074] In one embodiment, step b) is carried out in a solvent. In one embodiment, at least one solvent is a polar aprotic solvent. In one embodiment, the one or more solvents are selected from ethyl acetate (EtOAc), tetrahydrofuran, acetone, N,N-dimethylformamide, acetonitrile, methyl THF, NMP, DMA, and dimethyl sulfoxide. In one embodiment, step b) is carried out in EtOAc.

[0075] In one embodiment, reacting a compound of formula IV with a compound of formula V further comprises reacting with a base. In one embodiment, the base is selected from trimethylamine, sodium hydroxide, DIPEA, NMM, and potassium hydroxide. In one embodiment, the base is a tertiary amine. In one embodiment, the base is trimethylamine.

[0076] In one embodiment, step b) is performed between about 40°C and about 60°C. In one embodiment, the process is performed between about 45°C and about 60°C or between about 50°C and about 60°C. In one embodiment, step b) is performed between about 40°C and about 55°C, between about 40°C and about 50°C, or between about 40°C and about 45°C. In one embodiment, step b) is performed between about 45°C and about 55°C. In one embodiment, step d) is performed at about 40°C, about 45°C, about 50°C, about 55°C, or about 60°C. In one embodiment, step b) is performed below about 60°C. In one embodiment, step b) is performed at about 50°C.

[0077] In one embodiment, the compound of formula VI is separated in crystalline solid form. In one embodiment, step a) does not require chromatography for separation or purification. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula VI, based on liquid chromatographic area percentage (LCAP), is between about 95% and 99%. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula VI, based on LCAP, is at least about 95%.

[0078] In one embodiment, the crystalline compound of formula VI is produced from step b) in a yield between about 70% and 99%. In another embodiment, the compound of formula VI is produced in a yield between about 70% and 95%, about 70% and 90%, about 70% and 85%, about 70% and 80%, or about 70% and 75%. In yet another embodiment, the compound of formula VI is produced from step b) in a yield between about 75% and about 99%, about 80% and about 99%, about 85% and about 99%, about 90% and about 99%, or about 95% and about 99%. In yet another embodiment, the compound of formula VI is produced from step b) in a yield of about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 99%. In still another embodiment, the compound of formula VI is produced in a yield of about 85%. In yet another embodiment, the compound of formula VI is produced in approximately 90% yield.

[0079] In one embodiment, the compound of formula VI is generated from step b) in a yield of at least about 65%, at least about 70%, at least about 75%, at least about 80%, or at least about 85%. In another embodiment, formula VI is generated from step b) in a yield of at least about 80%.

[0080] In one embodiment, step b) is an amide coupling of the carboxylic acid of N,N-dimethylglycine with the amine of the compound of formula V to provide the compound of formula VI.

[0081] Step c

[0082] c) Reacting the compound of formula VI under coupling conditions to form the compound of formula I.

[0083]

[0084] Where a is an integer from 8 to 14;

[0085] b is an integer from 1 to 3; and

[0086] Z is the counter ion.

[0087] In one implementation, a and b are as described in step a). In one implementation, a is 12 and b is 2.

[0088] In one implementation, Z is Cl - or Br - In another implementation, Z is Br - .

[0089] In one embodiment, step c) is carried out in a solvent. In one embodiment, the solvent is a combination of solvents. In another embodiment, at least one solvent is polar aprotic. In another embodiment, all solvents are polar aprotic. In another embodiment, the one or more solvents are selected from EtOAc, N-methylpyrrolidone, tetrahydrofuran, methyl THF, dimethylacetamide, acetone, dimethylformamide, and acetonitrile. In one embodiment, step c) is carried out in ethyl acetate and acetonitrile.

[0090] In one embodiment, the coupling conditions include a reaction with a base. In one embodiment, the base is selected from sodium hydroxide, potassium hydroxide, DIPEA, NMM, and potassium carbonate. In one embodiment, the base is potassium carbonate.

[0091] It was found that residual water caused side reactions that affected purity and yield. Therefore, in one embodiment, step c) is carried out in a reaction vessel with an inert environment.

[0092] In one embodiment, step c) is performed between about 60°C and about 80°C. In one embodiment, the process is performed between about 65°C and about 80°C, or between about 70°C and about 80°C. In one embodiment, step c) is performed between about 60°C and about 75°C, or between about 60°C and about 70°C. In one embodiment, step c) is performed between about 65°C and about 75°C. In one embodiment, step c) is performed at about 60°C, about 65°C, about 70°C, about 75°C, or about 80°C. In one embodiment, step c) is performed at about 70°C.

[0093] In one embodiment, the compound of Formula I is produced from step c) in a yield between about 70% and 99%. In another embodiment, the compound of Formula I is produced in a yield between about 70% and 95%, about 70% and 90%, about 70% and 85%, about 70% and 80%, or about 70% and 75%. In yet another embodiment, the compound of Formula I is produced from step c) in a yield between about 75% and about 99%, about 80% and about 99%, about 85% and about 99%, about 90% and about 99%, or about 95% and about 99%. In yet another embodiment, the compound of Formula I is produced from step c) in a yield of about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 99%. In still another embodiment, the compound of Formula I is produced in a yield of about 90%. In yet another embodiment, the compound of formula I is produced in approximately 95% yield.

[0094] In one embodiment, the compound of formula I is generated from step c) in at least about 65% yield, at least about 70% yield, at least about 75% yield, at least about 80% yield, at least about 85% yield, or from step c) in at least about 90% yield or at least about 95% yield. In another embodiment, formula I is generated from step c) in at least about 90% yield.

[0095] In one embodiment, the compound of formula I is separated in crystalline solid form. In one embodiment, step a) does not require chromatography for separation or purification. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula I, based on LCAP, is between about 95% and 99.9%. In one embodiment, without chromatographic purification, the purity of the resulting compound of formula VI, based on LCAP, is at least about 95%.

[0096] In one embodiment, step c) is a coupling reaction. In one embodiment, step c) is the quaternization of the tertiary amine of the compound of formula VI with bromoethanol to give the bromide salt of formula I.

[0097] Overall process

[0098] In one embodiment, the compound of Formula I is produced from the compound of Formula II in a total yield between about 40% and about 80%. In one embodiment, the compound of Formula I is produced from the compound of Formula II in a yield between about 40% and about 70%, about 40% and about 60%, about 40% and about 50%, or about 40% and about 45%. In one embodiment, the compound of Formula I is produced from the compound of Formula II in a yield between about 50% and about 80%, about 60% and about 80%, about 70% and about 80%, or about 75% and about 80%. In one embodiment, the compound of Formula I is produced from the compound of Formula II in a yield of about 60%. In one embodiment, the compound of Formula I is produced from the compound of Formula II in a yield of about 65%. In one embodiment, the compound of formula I is generated from the compound of formula II in a yield of at least about 40%, at least about 50%, at least about 60%, or at least about 65%. In one embodiment, the compound of formula I is generated from the compound of formula II in a yield of at least about 50%. In one embodiment, the compound of formula I is generated from the compound of formula II in a yield of at least about 60%.

[0099] Example

[0100] Example 1. Synthesis of aziridine dibis(ethane-2,1-diyl)bistetradecanoate methanesulfonate methanesulfonic acid solvate

[0101]

[0102] Toluene (26.1 kg, 30.0 L, 30 L / kg) was charged into reactor 1, followed by N-Boc-diethanolamine (1.00 kg, 0.920 L). Observation: The presence of water may cause the myristylation reaction to stop and result in higher levels of impurities (myristic acid). The reactor should be thoroughly rinsed and dried before use. The jacket temperature was set to 15°C–25°C (target 20°C). N-methylmorpholine (1.68 kg, 1.82 L, 3.4 equivalents) was charged. Myristoyl chloride (2.65 kg, 2.94 L, 2.2 equivalents) was charged into the reactor after at least 30 minutes, maintaining the batch temperature <30°C (target 20°C). The reaction was aged at 15°C–25°C (target 20°C) for at least 5 hours.

[0103] Add 1N acetic acid (11.0 kg, 10 L, 10 L / kg) in a semi-saline solution to the reactor. Heat the reactants to 25°C–35°C (target 30°C) with stirring and age for 30 minutes. Stop stirring and allow the reaction to allow the phases to settle for at least 30 minutes. Separate the phases and send the bottom aqueous phase to the waste. Repeat this washing process.

[0104] The organic layer was distilled under vacuum (P < 100 mbar) to a final volume of 14 L / kg, maintaining the jacket setpoint temperature at 45°C. Observation: Distillation is typically performed using a put-and-take method; it is recommended to add a minimum volume of 20 L / kg of toluene to dry the stream. In subsequent steps, the presence of water may cause product decomposition. These impurities significantly increase product solubility, which in turn significantly reduces the yield. Furthermore, changes in solubility affect the seed point in step 28.

[0105] The reaction mixture was transferred to reactor 2 via a fine filter to remove precipitated NaCl. Toluene was loaded into reactor 1 as a vessel rinse solution using a spray nozzle and transferred to reactor 2 via a fine filter line (3.48 kg, 4.0 L, 4.0 L / kg). The reaction was heated to 30°C–40°C (target 35°C) under stirring and a nitrogen atmosphere. Methanesulfonic acid (0.936 kg, 0.693 L, 2.0 equivalent) was added after at least 30 minutes, maintaining the batch temperature at 30°C–40°C (target 35°C). Note: Anhydrous methanesulfonic acid (KF < 0.75 wt%) was used. Excess water may cause decomposition of BMT-334112. Anhydrous ethyl acetate (2.70 kg, 3.0 L, 3.0 L / kg) was added to the reaction mixture, and the reaction was aged at 30°C–40°C (target 35°C) for at least 1 hour, then cooled to 27°C–31°C (target 29°C).

[0106] 0.050 kg (5 wt%) of aziridine dibis(ethane-2,1-diyl)bistetradecanoate methanesulfonate methanesulfonic acid solvation seed was added to the reaction vessel. The reaction was then cooled to 10°C-20°C (target 15°C) for at least five hours.

[0107] Achieve the following cooling ramp:

[0108] i. After 30% of the total cooling time, from 29°C to 27.25°C.

[0109] ii. After 30% of the total cooling time, from 27.25°C to 24.5°C.

[0110] iii. After 40% of the total cooling time, from 24.5°C to 15°C.

[0111] Observation: Due to the effect of temperature on the solubility of the product in toluene / ethyl acetate, nonlinear cooling is implemented to facilitate filtration rate and impurity removal.

[0112] The reaction batch was then cooled at 10°C–20°C (target 15°C) for 1 hour and filtered. Tert-butyl methyl ether (MTBE, 2.22 kg, 3.0 L, 3.0 L / kg) was added to reactor 1 as crystallizer wash and cooled to 10°C–20°C (target 15°C). The product filter cake was then washed with MTBE wash from reactor 1, and the wash was filtered. The filter cake was dried under vacuum and nitrogen purging at a jacket setpoint of 20°C–30°C (target 25°C) to produce crystalline alkyldimethylbis(ethane-2,1-diyl)bistetradecanoate methanesulfonate methanesulfonic acid solvate in 90% yield. Observation: Increased temperature may cause the wet filter cake to melt / dissolve and lead to significant product degradation.

[0113] Example 2: Synthesis of ((dimethylglycyl)azanidinediyl)bis(ethane-2,1-diyl)bistetradecanoate oxalate

[0114]

[0115] Charge EtOAc into reactor 3. Dry EtOAc (KF < 200 ppm) should be used to minimize the decomposition of the neopentanoyl chloride (PivCl) reagent. Charge N,N-dimethylglycine (0.20 kg, 1.4 equivalents) into reactor 3 to produce a slurry. The presence of water may cause incomplete reaction. Before use, thoroughly flush the reactor with EtOAc. Then charge triethylamine (TEA) (0.62 kg, 4.4 equivalents) into reactor 3, followed by PivCl (0.32 kg, 1.9 equivalents). Run the reaction at 50°C for at least 3 h, then cool to 20°C. Charge aziridine dimethyl bis(ethane-2,1-dimethyl)bistetradecanoate (1 kg, 1 equivalent) into reactor 3, followed by EtOAc (6 L, 6 L / kg). Run the reaction at 20°C for 1.5 h.

[0116] Add water (8.32 L, 8.32 L / kg) to reactor 3. The water will remove organic salts formed as reaction byproducts. Add 25% sodium chloride brine (1.68 L, 1.68 L / kg) to reactor 3 and age at 20°C for at least 0.5 h. Stop stirring and allow the reaction to stand at 20°C for at least 0.5 h. Separate the phases and send the lower aqueous layer from reactor 3 to waste. Add carbonate buffer pH (10 L, 10 L / kg) to reactor 3 and age at 20°C for at least 1 h. Stop stirring and allow the reaction to stand at 20°C for at least 1 h. Separate the phases and send the lower aqueous layer from reactor 3 to waste.

[0117] EtOAc (5 L, 5.0 L / kg) was then loaded into reactor 3 and cooled to 5°C. The batch was then distilled to a concentration of 10 L / kg. Distillation should be carried out at a pressure of 50 mbar, with the jacket temperature setpoint between 25°C and 35°C. Temperatures above 35°C may lead to the formation of impurities over prolonged periods.

[0118] Toluene (10 L, 10.0 L / kg) was added to reactor 3, and the reaction was heated to 52°C. After at least 1 hour, oxalic acid solution (5 L, 5.0 L / kg, 1.1 equivalents) was added above the surface of reactor 3, and the mixture was aged for 1 hour. The batch was then cooled according to the following plan:

[0119] a. The batch of material was cooled to 46°C after 3.5 hours and aged for 2 hours.

[0120] b. Cool the batch to 40°C over 2.5 hours.

[0121] c. Cool the batch to 20°C for at least 2 hours and age for 1 hour.

[0122] The resulting slurry was filtered and EtOAc (2 L, 2.0 L / kg) was charged into reactor 3. The filter cake was washed using this EtOAc (2.0 L / kg). The filter cake was then dried under vacuum at a jacket setpoint temperature ≤55°C (target 50°C–55°C) to produce ((dimethylglycyl)azanyl)bis(ethane-2,1-diyl)bistetradecanoate oxalate in 94% yield.

[0123] Example 3: N 1 N 19 Synthesis of bis((S)-16,20-diamino-15-oxo-4,7,10-trioxa-14-azaeicosyl)-4,7,10,13,16-pentaoxa-nonadecanidamide (HEDC).

[0124]

[0125] Add 20 L of EtOAc to reactor 1, followed by 1 kg of ((dimethylglycyl)azanyl)bis(ethane-2,1-diyl)bistetradecanoate oxalate. Add 15 L of a pH 9.5 carbonate buffer solution to reactor 1 and heat the reaction to 30 °C, then age for at least 1 h. Cool the reaction to 20 °C, then stop stirring and allow the reaction to stand at 20 °C for at least 1 h. Separate the phases, and send the lower aqueous layer from reactor 1 to the waste. Add 9.16 L of water to reactor 1, followed by 0.84 L of 25% sodium chloride brine. Age the reaction at 20 °C for at least 0.5 h with stirring. Stop stirring and allow the reaction to stand at 20 °C for at least 0.5 h. Then separate the phases, and send the lower aqueous layer from reactor 1 to the waste.

[0126] The reaction mixture from reactor 1 was then filtered into reactor 2. Reactor 1 was then flushed with EtOAc (5 L) and transferred to reactor 2 through a fine filter. The solution in reactor 2 was then cooled to 5 °C and concentrated to 5 ± 0.5 L / kg by distillation at 50 mbar under a jacket temperature setpoint between 25 °C and 35 °C. 2-Bromoethanol (0.40 kg) was added to reactor 2. Then MeCN (6 L) was added. The solution in reactor 2 was then cooled to 5 °C and concentrated to 5 ± 0.5 L / kg by distillation at 50 mbar under a jacket temperature setpoint between 25 °C and 35 °C. Reactor 2 was then heated to 70 °C and aged for at least 24 h, and then cooled to 60 °C.

[0127] EtOAc was loaded into reactor 2 (15L) and cooled to 45°C. HEDC seeds (0.005kg) were added to reactor 2 and aged at 45°C for at least 1 hour. Reactor 2 was then cooled according to the following schedule:

[0128] a. Cool to 40°C after 3 hours.

[0129] b. Cool to 35°C in 1.5 hours.

[0130] c. Cool to 30°C in 1 hour.

[0131] d. Cool to 20°C in 1 hour.

[0132] e. Aging at 20°C for at least 1 hour.

[0133] The resulting slurry was then filtered. Reactor 3 was flushed with EtOAc. The filter cake was then flushed with the same EtOAc. The filter cake was then thoroughly washed with EtOAc at least twice (4 L each time). The final product filter cake was dried under vacuum at a jacket setpoint temperature ≤55°C (target 50°C-55°C) to produce HEDC in 92% yield.

Claims

1. A method for synthesizing compounds of formula I. Where a is an integer between 8 and 14; b is an integer between 1 and 3; and Z is Cl - or Br - , The method includes a) Make the compound of formula II Where R is a protecting group; Compounds of Formula III Where X is a halogen; The reaction forms a compound of formula IV. b) Mixing the compound of formula IV with the compound of formula V The reaction forms a compound of formula VI. as well as c) Reacting the compound of formula VI with bromoethanol under coupling conditions to form the compound of formula I.

2. The method according to claim 1, wherein a is 12.

3. The method according to claim 1, wherein b is 2.

4. The method according to claim 1, wherein R is selected from carboxybenzyl, p-methoxybenzylcarbonyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, acetyl, trifluoroacetyl, benzoyl, benzyl, carbamate, p-methoxybenzyl, 3,4-dimethoxybenzyl, toluenesulfonyl, trichloroethyl chloroformate, (4-nitrophenyl)sulfonyl, methyl, ethyl, propyl, n-butyl, tert-butyl, succinimide, 2,6-dimethylphenol, 2,6-diisopropylphenol, 2,6-ditert-butylphenol, trimethylsilyl, allyl, 1,1-dimethylallyl, 2,2,2-trifluoroethyl, phenyl, and 4-methoxybenzyl.

5. The method according to claim 4, wherein R is tert-butoxycarbonyl.

6. The method according to claim 1, wherein X is Cl or Br.

7. The method according to claim 6, wherein X is Cl.

8. The method according to claim 1, wherein Z is Cl - .

9. The method of claim 1, wherein Z is Br - .

10. The method of claim 1, wherein the yield of step a) is at least 80%.

11. The method of claim 1, wherein the yield of step b) is at least 80%.

12. The method according to claim 1, wherein in step c), bromoethanol is used between 2 and 4 equivalents, and the compound of formula VI is used in 1 equivalent.

13. The method of claim 12, wherein in step c), bromoethanol is used in an amount of 2.2 equivalents.

14. The method of claim 1, wherein the compound of formula IV is isolated in solid form.

15. The method of claim 14, wherein the compound of formula IV is isolated in crystalline solid form.

16. The method of claim 1, wherein the compound of formula VI is isolated in crystalline solid form.

17. The method according to claim 1, wherein the compound of formula I is 。 18. The method of claim 1, wherein the compound of formula I is produced by the compound of formula II in a yield of at least 60%.

Images