Compound serving as intermediate for production of amino lipid or salt thereof, and amino lipid compound production method using same

Abstract

The present invention addresses the problem of providing: an amino lipid compound production method by which it becomes possible to synthesize a carbamate with high yield; and an intermediate compound for use in the amino lipid compound production method. The present invention provides a compound represented by formula (1). In the formula, R1 represents a hydrogen atom or a hydrocarbon group having 1 to 12 carbon atoms; R2 represents a divalent hydrocarbon group having 1 to 12 carbon atoms; R3 represents a hydrocarbon group having 1 to 24 carbon atoms; and X1 represents (AA) or a halogen atom.

Description

Compound as an intermediate for the production of amino lipid or its salt, and method for producing amino lipid compound using the same

[0001] The present invention relates to a compound as an intermediate for the production of an amino lipid or a salt thereof, and a method for producing an amino lipid compound using the compound.

[0002] Nucleic acid drugs have a clear mechanism of action against diseases and few side effects, and are expected to be next-generation pharmaceuticals, and their development is actively underway. One known technique for delivering nucleic acids to cells is to administer nucleic acids encapsulated in liposomes or lipid particles. This technique uses lipids with substituents that become cationic at low pH, such as amino groups, and achieves nucleic acid delivery by imparting an appropriate charge to the particles.

[0003] As compounds to be contained in lipid particles, Patent Document 1 describes an amino lipid synthesized by condensation of an imidazole carboxylic acid ester with an alcohol, and Patent Document 2 describes an amino lipid synthesized by condensation of a 1-triazole carboxylic acid ester intermediate with an alcohol.

[0004] On the other hand, Patent Document 3 describes the use of triazolecarboxylic acid amides as agricultural chemicals and fungicides, and Non-Patent Document 1 describes the use of triazolecarboxylic acid amides as medicines.

[0005] International Publication No. WO2022 / 230964 International Publication No. WO2024 / 014430 EP0234255B1

[0006] J. Med. Chem. 2024,67,3,1758.

[0007] An object of the present invention is to provide a method for producing an amino lipid compound, which can synthesize carbamates in high yield, as well as an intermediate compound used in the method for producing the amino lipid compound.

[0008] As a result of extensive research aimed at solving the above-mentioned problems, the present inventors have discovered that carbamates can be synthesized in high yields by using carboxamides (carboxylic acid amides) as production intermediates. Unlike the case of using imidazole carboxylic acid esters, imidazole carboxylic acid amides have low reactivity and are not susceptible to nucleophilic attack by alcohols. The addition of alkylamines or DMAP (N,N-dimethylaminopyridine) was investigated to promote the reaction, but the reaction progress rate was low. It is presumed that complete abstraction of the protons from the alcohol reactant to convert it into an alkoxide is important for the reaction to proceed. The present invention was completed based on the above findings. According to the present invention, the following inventions are provided.

[0009] <1> A compound represented by formula (1). In the formula, R １ represents a hydrogen atom or a hydrocarbon group having 1 to 12 carbon atoms; R ２ represents a divalent hydrocarbon group having 1 to 12 carbon atoms, and R ３ represents a hydrocarbon group having 1 to 24 carbon atoms; X １ teeth, or a halogen atom. １ represents a hydrocarbon group having 3 to 8 carbon atoms; R ２ represents a divalent hydrocarbon group having 1 to 5 carbon atoms; R ３ represents a hydrocarbon group having 12 to 24 carbon atoms. １ The compound according to <1>, wherein the halogen atom represented by is a chlorine atom. <4> or <5> A method for producing a compound represented by formula (3) or a salt thereof, comprising reacting the compound represented by formula (1) according to <1> with a compound represented by formula (2) or a salt thereof: In the formula, R４ and R ５ each independently represents a hydrocarbon group having 1 or 2 carbon atoms; R ４ and R ５ each independently represents an optionally substituted hydrocarbon group having 1 to 18 carbon atoms; R ４ and R ５ The substituents on the optionally substituted hydrocarbon group having 1 to 18 carbon atoms represented by the formula (I) are each independently —OH, —COOH, —NR ９ R １０ , -OC(O)OR １１ , -C(O)O-R １２ , —OC(O)—R １３ , -O-R １４ , —C(O)NR １５ R １６ , -NR １７ C(O)R １８ , -N(R １９ ) S (O) ２ R ２０ , -N(R ２１ )C(O)N(R ２２ ) R ２３ , -N(R ２４ ) C(S)N(R ２５ ) R ２６ , -OC(O)N(R ２７ ) R ２８ , or -N(R ２９ )C(O)OR ３０ indicates R ９ and R １０ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R １１ , R １２ , R １３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０each independently represents a hydrogen atom or an optionally substituted hydrocarbon group having 1 to 24 carbon atoms; R １１ , R １２ , R １３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０ The substituent on the optionally substituted hydrocarbon group having 1 to 24 carbon atoms represented by is an aryl group having 6 to 20 carbon atoms, a heterocyclic group, —OH, —COOH, or —NR ３１ R ３２ indicates R ３１ and R ３２ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R ４ and R ５ , or R ４ and R ６ may be joined together to form a 4- to 7-membered ring, R ６ represents a hydrocarbon group having 2 to 4 carbon atoms, and R ７ and R ８ each independently represents a hydrocarbon group having 2 or 3 carbon atoms, or R ５ and R ６ may be linked to form a cyclic structure, In the formula, R １ , R ２ and R ３ is the same as defined in <1>, R ４ , R ５ , R ６ , R ７ and R ８ is defined as in formula (2).

[0010] According to the present invention, amino lipids can be produced in high yield.

[0011] The present invention will be described in detail below. In this specification, the symbol "to" indicates a range that includes the numerical values ​​before and after it as the minimum and maximum values, respectively.

[0012] <Intermediate for producing amino lipid or its salt> The intermediate of the present invention is a compound represented by formula (1). In the formula, R １ represents a hydrogen atom or a hydrocarbon group having 1 to 12 carbon atoms; R ２ represents a divalent hydrocarbon group having 1 to 12 carbon atoms, and R ３ represents a hydrocarbon group having 1 to 24 carbon atoms; X １ teeth, or a halogen atom.

[0013] R １ R preferably represents a hydrocarbon group having 2 to 10 carbon atoms, more preferably a hydrocarbon group having 3 to 8 carbon atoms, even more preferably a hydrocarbon group having 5 to 8 carbon atoms, and particularly preferably a hydrocarbon group having 6 or 7 carbon atoms. ２ represents a divalent hydrocarbon group preferably having 1 to 10 carbon atoms, more preferably a divalent hydrocarbon group having 1 to 8 carbon atoms, even more preferably a divalent hydrocarbon group having 1 to 5 carbon atoms, and particularly preferably a hydrocarbon group having 5 carbon atoms. ３ represents a hydrocarbon group having preferably 4 to 24 carbon atoms, more preferably a hydrocarbon group having 8 to 24 carbon atoms, and even more preferably a hydrocarbon group having 12 to 24 carbon atoms. １ The halogen atom represented by the formula (I) includes a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, etc., and is preferably a chlorine atom.

[0014] Specific examples of the compound represented by formula (1) are shown below, but the invention is not limited to these.

[0015] R １ The hydrocarbon group having 1 to 12 carbon atoms represented by is preferably an alkyl group having 1 to 12 carbon atoms, an alkenyl group having 2 to 12 carbon atoms, or an alkynyl group having 2 to 12 carbon atoms, and more preferably an alkyl group having 1 to 12 carbon atoms. The hydrocarbon group having 1 to 12 carbon atoms may be linear or branched, and may be linear or cyclic. Specific examples of the hydrocarbon group having 1 to 12 carbon atoms include a methyl group, an ethyl group, a propyl group, an isopropyl group, a cyclopropyl group, a butyl group, an isobutyl group, a tert-butyl group, a cyclobutyl group, a pentyl group, a cyclopentyl group, a hexyl group, a cyclohexyl group, a heptyl group, an octyl group, a nonyl group, a decyl group, an undecyl group, and a dodecyl group.

[0016] R ２ The divalent hydrocarbon group having 1 to 12 carbon atoms represented by is preferably an alkylene group having 1 to 12 carbon atoms or an alkenylene group having 2 to 12 carbon atoms. The divalent hydrocarbon group having 1 to 12 carbon atoms may be linear or branched, and may be linear or cyclic. Specific examples of the divalent hydrocarbon group having 1 to 12 carbon atoms include a methylene group, an ethylene group, a trimethylene group, a tetramethylene group, a pentamethylene group, a hexamethylene group, a heptamethylene group, an octamethylene group, a nonamethylene group, a decamethylene group, an undecamethylene group, and a dodecamethylene group.

[0017] R ３The hydrocarbon group having 1 to 24 carbon atoms represented by is preferably an alkyl group, an alkenyl group, or an alkynyl group. The alkyl group may be linear or branched, and may be chain-like or cyclic. Specific examples of the alkyl group include a methyl group, an ethyl group, a propyl group, an isopropyl group, a cyclopropyl group, a butyl group, an isobutyl group, a tert-butyl group, a cyclobutyl group, a pentyl group, a cyclopentyl group, a hexyl group, a cyclohexyl group, a heptyl group, an octyl group, a nonyl group, a decyl group, an undecyl group, a dodecyl group, a tridecyl group, a trimethyldodecyl group (preferably a 3,7,11-trimethyldodecyl group), a tetradecyl group, a pentadecyl group, a hexadecyl group, a tetramethylhexadecyl group (preferably a 3,7,11,15-tetramethylhexadecyl group), a heptadecyl group, an octadecyl group, a 2-butylhexyl group, and a 2-butyloctyl group. , 1-pentylhexyl group, 2-pentylheptyl group, 3-pentyloctyl group, 1-hexylheptyl group, 1-hexylnonyl group, 2-hexyloctyl group, 2-hexyldecyl group, 3-hexylnonyl group, 1-heptyloctyl group, 2-heptylnonyl group, 2-heptylundecyl group, 3-heptyldecyl group, 1-octylnonyl group, 2-octyldecyl group, 2-octyldodecyl group, 3-octylundecyl group, 2-nonylundecyl group, 3-nonyldodecyl group, 2-decyldodecyl group, 2-decyltetradecyl group, 3-decyltridecyl group, 2-(4,4-dimethylpentan-2-yl)-5,7,7-trimethyloctyl group, and the like.

[0018] The alkenyl group may be linear or branched, linear or cyclic. Specific examples include allyl, prenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl (preferably, (Z)-2-nonenyl or (E)-2-nonenyl), decenyl, undecenyl, dodecenyl, dodecadienyl, tridecenyl (preferably, (Z)-tridec-8-enyl), tetradecenyl (preferably, tetradec-9-enyl), and pentadecenyl (preferably, (Z)-pentadecen-8-enyl). , a hexadecenyl group (preferably a (Z)-hexadec-9-enyl group), a hexadecadienyl group, a heptadecenyl group (preferably a (Z)-heptadeca-8-enyl group), a heptadecadienyl group (preferably a (8Z,11Z)-heptadeca-8,11-dienyl group), an octadecenyl group (preferably a (Z)-octadec-9-enyl group), an octadecadienyl group (preferably a (9Z,12Z)-octadeca-9,12-dienyl group), and the like.

[0019] The alkynyl group may be linear or branched, open-chain or cyclic, and specific examples thereof include a propargyl group, a butynyl group, a pentynyl group, a hexynyl group, a heptynyl group, an octynyl group, a nonynyl group, a decynyl group, an undecynyl group, a dodecynyl group, a tetradecynyl group, a pentadecynyl group, a hexadecynyl group, a heptadecynyl group, and an octadecynyl group.

[0020] Preferably, all of the above alkenyl groups have one or two double bonds, and preferably, all of the alkynyl groups have one or two triple bonds.

[0021] <Method for producing an amino lipid compound represented by formula (3) or a salt thereof> The present invention relates to a method for producing a compound represented by formula (3) or a salt thereof, which comprises reacting the compound represented by formula (1) described above with a compound represented by formula (2) or a salt thereof. In the formula, R ４ and R ５each independently represent a hydrocarbon group having 1 to 18 carbon atoms, R4 and R5 each independently represent a hydrocarbon group having 1 to 18 carbon atoms which may be substituted, and the substituents on the hydrocarbon group having 1 to 18 carbon atoms which may be substituted represented by R4 and R5 each independently represent -OH, -COOH, -NR ９ R １０ , -OC(O)OR １１ , -C(O)O-R １２ , —OC(O)—R １３ , -O-R １４ , —C(O)NR １５ R １６ , -NR １７ C(O)R １８ , -N(R １９ ) S (O) ２ R ２０ , -N(R ２１ )C(O)N(R ２２ ) R ２３ , -N(R ２４ ) C(S)N(R ２５ ) R ２６ , -OC(O)N(R ２７ ) R ２８ , or -N(R ２９ )C(O)OR ３０ indicates R ９ and R １０ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R １１ , R １２ , R １３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０ each independently represents a hydrogen atom or an optionally substituted hydrocarbon group having 1 to 24 carbon atoms; R １１ , R １２ , R１３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０ The substituent on the optionally substituted hydrocarbon group having 1 to 24 carbon atoms represented by is an aryl group having 6 to 20 carbon atoms, a heterocyclic group, —OH, —COOH, or —NR ３１ R ３２ indicates R ３１ and R ３２ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R ４ and R ５ , or R ４ and R ６ may be joined together to form a 4- to 7-membered ring, R ６ represents a hydrocarbon group having 2 to 4 carbon atoms, and R ７ and R ８ each independently represents a hydrocarbon group having 2 or 3 carbon atoms, or R ５ and R ６ may be linked to form a cyclic structure, In the formula, R １ , R ２ and R ３ is defined as in formula (1), and R ４ , R ５ , R ６ , R ７ and R ８ is defined as in formula (2).

[0022] R ４ and R ５ The hydrocarbon group having 1 or 2 carbon atoms represented by R is preferably an alkyl group, more preferably a methyl group or an ethyl group, and even more preferably an ethyl group. ６The hydrocarbon group having 2 to 4 carbon atoms represented by is preferably an alkylene group or an alkenylene group, and more preferably an alkylene group. Specific examples of the hydrocarbon group having 2 to 4 carbon atoms include an ethylene group, a trimethylene group, and a tetramethylene group, and an ethylene group is preferred. ７ and R ８ The hydrocarbon group having 2 or 3 carbon atoms represented by is preferably an alkylene group or an alkenylene group, more preferably an alkylene group. Specific examples of the hydrocarbon group having 2 or 3 carbon atoms include an ethylene group or a trimethylene group, with an ethylene group being preferred.

[0023] The compound represented by formula (2) may form a salt. The compound represented by formula (3) may form a salt. Examples of salts of basic groups include salts with mineral acids such as hydrochloric acid, hydrobromic acid, nitric acid, and sulfuric acid; salts with organic carboxylic acids such as formic acid, acetic acid, citric acid, oxalic acid, fumaric acid, maleic acid, succinic acid, malic acid, tartaric acid, aspartic acid, trichloroacetic acid, and trifluoroacetic acid; and salts with sulfonic acids such as methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, mesitylenesulfonic acid, and naphthalenesulfonic acid. Of the above-mentioned salts, preferred salts include pharmacologically acceptable salts.

[0024] <Production Method> The production method of the compound of the present invention will be described. The compound of the present invention can be produced by combining known methods, and can be produced, for example, according to the production method shown below.

[0025] [Production Method 1] A method for producing a compound of formula [1] from compounds of formula [2] and formula [3].

[0026] In the formula, R １ , R ２ , R ３ , R ４ , R ５ , R ６ , R ７ and R ８ has the same meaning as above; X １ means a leaving group.

[0027] Examples of leaving groups include a chloro group, a fluoro group, a bromo group, a trichloromethoxy group, a 4-nitro-phenoxy group, a 2,4-dinitrophenoxy group, a 2,4,6-trichlorophenoxy group, a pentafluorophenoxy group, a 2,3,5,6-tetrafluorophenoxy group, an imidazolyl group, a triazolyl group, a 3,5-dioxo-4-methyl-1,2,4-oxadiazolidyl group, and an N-hydroxysuccinimidyl group.

[0028] The compound of formula [1] can be produced by reacting a compound of formula [2] with a compound of formula [3] in the presence of a base. For example, 2,2'-((3-(diethylamino)propyl)azanediyl)bis(ethan-1-ol) is known as a compound of formula [3], and it can be synthesized according to the examples described in WO 2022 / 230964. The solvent used in this reaction is not particularly limited as long as it does not affect the reaction, and examples include halogenated hydrocarbons, ethers, esters, amides, nitriles, sulfoxides, and aromatic hydrocarbons. These solvents may be used in combination. Preferred solvents include ethers, and tetrahydrofuran is more preferred. The amount of solvent used is not particularly limited, and may be 1 to 500 times (v / w) the amount of the compound of formula [2]. The base used in this reaction includes an inorganic base or an organic base. Specific examples include potassium carbonate, sodium carbonate, lithium carbonate, potassium phosphate, sodium phosphate, lithium phosphate, sodium hydride, triethylamine, N,N-diisopropylethylamine, 4-methylmorpholine, pyridine, 1,8-diazabicyclo[5.4.0]-7-undecene, and N,N-dimethylaminopyridine, with sodium hydride and potassium tert-butoxide being particularly preferred. The amount of base used may be 1 to 50 times, preferably 1 to 10 times, the molar ratio of the compound of formula [3]. The amount of the compound of formula [3] used is not particularly limited, but may be 0.1 to 10 times the molar ratio of the compound of formula [2]. This reaction may be carried out at -30 to 150°C, preferably 0 to 100°C, for 5 minutes to 48 hours.

[0029] [Production Method 2] A method for producing a compound of formula [2] from a compound of formula [4].

[0030] In the formula, R １ , R ２ and R ３ has the same meaning as above; X １ and X ２ means the same leaving group as above.

[0031] The compound of formula [2] can be produced by reacting a compound of formula [5] with a compound of formula [4] in the presence or absence of a base. Known examples of compounds of formula [5] include 1,1'-carbonyldi(1,2,4-triazole), 1,1'-carbonyldiimidazole, 4-nitrophenyl chloroformate, triphosgene, and phosgene. The solvent used in this reaction is not particularly limited as long as it does not affect the reaction, and examples include halogenated hydrocarbons, ethers, esters, amides, nitriles, sulfoxides, and aromatic hydrocarbons. These solvents may be used in combination. Preferred solvents include ethers, with tetrahydrofuran being more preferred. The amount of solvent used is not particularly limited, and may be 1 to 500 times (v / w) the amount of the compound of formula [4]. The base used in this reaction includes inorganic bases and organic bases. The base is preferably an organic base, and specific examples include triethylamine, N,N-diisopropylethylamine, 4-methylmorpholine, pyridine, 1,8-diazabicyclo[5.4.0]-7-undecene, and N,N-dimethylaminopyridine. The amount of the base used may be 0 to 50 times, preferably 0 to 10 times, the molar amount of the compound of formula [4]. The amount of the compound of formula [5] used is not particularly limited, but may be 1 to 10 times the molar amount of the compound of formula [4]. This reaction may be carried out at -30 to 150°C, preferably 0 to 100°C, for 5 minutes to 48 hours.

[0032] [Production Method 3] A method for producing a compound of formula [4] from a compound of formula [6].

[0033] In the formula, R １ , R ２ and R ３ has the same meaning as above; X ３ means the same leaving group as above.

[0034] The compound of formula [4] can be produced by reacting a compound of formula [6] with a compound of formula [7] in the presence or absence of a base and in the presence or absence of an additive. Examples of compounds of formula [7] include 1-heptylamine and 1-hexylamine. The solvent used in this reaction is not particularly limited as long as it does not affect the reaction, and examples include halogenated hydrocarbons, ethers, esters, amides, nitriles, sulfoxides, and aromatic hydrocarbons. These solvents may be used in mixtures. Preferred solvents include mixed solvents of amides and esters, and a mixed solvent of dimethylformamide and ethyl acetate is more preferred. The amount of solvent used is not particularly limited, and may be 1 to 500 times (v / w) the amount of the compound of formula [6]. The base used in this reaction includes inorganic bases and organic bases. Specific examples of the base include potassium hydroxide, sodium hydroxide, lithium hydroxide, potassium carbonate, sodium carbonate, lithium carbonate, potassium phosphate, sodium phosphate, lithium phosphate, triethylamine, N,N-diisopropylethylamine, 4-methylmorpholine, pyridine, 1,8-diazabicyclo[5.4.0]-7-undecene, and N,N-dimethylaminopyridine, with potassium carbonate being more preferred. The amount of the base used may be 1 to 50 times, preferably 1 to 10 times, the molar ratio of the compound of formula [6]. The amount of the compound of formula [7] used is not particularly limited, but may be 1 to 10 times the molar ratio of the compound of formula [6]. Specific examples of additives used in this reaction include lithium iodide, sodium iodide, potassium iodide, benzyltriethylammonium iodide, and benzyltriethylammonium bromide. The amount of the additive used may be 0.1 to 10 times the molar ratio of the compound of formula [6]. This reaction may be carried out at a temperature of from -30 to 150°C, preferably from 0 to 100°C, for 5 minutes to 48 hours.

[0035] [Production Method 4] A method for producing a compound of formula [6].

[0036] In the formula, R ２ , R３ and X ３ has the same meaning as above; X means a hydroxyl group or a leaving group; the leaving group has the same meaning as above.

[0037] The compound of formula [6] can be produced by reacting a compound of formula [8] with a compound of formula [9] in the presence or absence of an acid, in the presence or absence of a condensing agent or acid halide, and in the presence or absence of a base. Known examples of compounds of formula [8] include 6-bromohexanoic acid and 5-bromovaleric acid. Known examples of compounds of formula [9] include 2-butyl-1-octanol, 2-pentyl-1-heptanol, and 1-decanol. The solvent used in this reaction is not particularly limited as long as it does not affect the reaction, and examples include halogenated hydrocarbons, ethers, esters, amides, nitriles, sulfoxides, and aromatic hydrocarbons. These solvents may be used in combination. Preferred solvents include aromatic hydrocarbons and ethers, with toluene and tetrahydrofuran being more preferred. The amount of solvent used is not particularly limited, but may be 1 to 500 times (v / w) the amount of the compound of formula [8]. The acid used in this reaction includes inorganic and organic acids. The acid is preferably a sulfonic acid, and specific examples include sulfuric acid, 4-toluenesulfonic acid, and methanesulfonic acid. Condensing agents used in this reaction include, for example, carbodiimides such as N,N'-dicyclohexylcarbodiimide and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide; carbonyls such as carbonyldiimidazole; acid azides such as diphenylphosphoryl azide; acid cyanides such as diethylphosphoryl cyanide; 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline; and uroniums such as O-benzotriazol-1-yl-1,1,3,3-tetramethyluronium hexafluorophosphate and O-(7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate.Examples of acid halides used in this reaction include carboxylic acid halides such as acetyl chloride and trifluoroacetyl chloride; sulfonic acid halides such as methanesulfonyl chloride and tosyl chloride; and chloroformates such as ethyl chloroformate and isobutyl chloroformate. Examples of bases used in this reaction include inorganic and organic bases. Organic bases are preferred, and specific examples include triethylamine, N,N-diisopropylethylamine, 4-methylmorpholine, pyridine, 1,8-diazabicyclo[5.4.0]-7-undecene, and N,N-dimethylaminopyridine. The amount of base used may be 1 to 50 times, preferably 1 to 10 times, the molar ratio of the compound of formula [8]. The amount of compound of formula [9] used is not particularly limited, but may be 0.8 to 10 times (v / w) the amount of compound of formula [8]. This reaction may be carried out at -30 to 150°C, preferably 0 to 100°C, for 5 minutes to 48 hours.

[0038] <Use of the compound represented by formula (3) or a salt thereof> The compound represented by formula (3) or a salt thereof produced by the production method of the present invention can be used to prepare lipid particles. When preparing lipid particles, in addition to the compound represented by formula (3) or a salt thereof, at least one lipid selected from the group consisting of sterols, neutral lipids, and lipids having nonionic hydrophilic polymer chains can be used. The lipid particles can further contain nucleic acids.

[0039] In the lipid particles, the amount of amino lipid blended is preferably 20 mol % to 80 mol %, more preferably 35 mol % to 70 mol %, and even more preferably 40 mol % to 65 mol %, relative to the total lipid amount.

[0040] Examples of sterols include, but are not limited to, cholesterol, phytosterols (sitosterol), stigmasterol, fucosterol, spinasterol, brassicasterol, ergosterol, cholestanone, cholestenone, coprostanol, cholesteryl-2'-hydroxyethyl ether, and cholesteryl-4'-hydroxybutyl ether. Of these, cholesterol is preferred. The amount of sterol blended is preferably 10 mol% to 60 mol%, more preferably 20 mol% to 55 mol%, and even more preferably 25 mol% to 50 mol%, based on the total lipid amount.

[0041] Neutral lipid is not particularly limited, but can be phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, ceramide etc., and preferably phosphatidylcholine and phosphatidylethanolamine.In addition, neutral lipid can be used alone or in combination with a plurality of different neutral lipids.In lipid particle, the amount of neutral lipid is preferably 3 mol% or more and 55 mol% or less with respect to the total amount of constituent lipid components.

[0042] Examples of nonionic hydrophilic polymers in lipids having nonionic hydrophilic polymer chains include, but are not limited to, nonionic vinyl polymers, nonionic polyamino acids, nonionic polyesters, nonionic polyethers, nonionic natural polymers, nonionic modified natural polymers, and block polymers or graft copolymers made up of two or more of these polymers as building blocks.Among these nonionic hydrophilic polymers, preferred are nonionic polyethers, nonionic polyesters, nonionic polyamino acids or nonionic synthetic polypeptides, more preferred are nonionic polyethers or nonionic polyesters, even more preferred are nonionic polyethers or nonionic monoalkoxy polyethers, and particularly preferred are polyethylene glycols (hereinafter, polyethylene glycols are also referred to as PEG).

[0043] Lipids having a nonionic hydrophilic polymer include, but are not limited to, PEG-modified phosphoethanolamine, diacylglycerol PEG derivatives, dialkylglycerol PEG derivatives, cholesterol PEG derivatives, and ceramide PEG derivatives. Among these, diacylglycerol PEG is preferred. The weight-average molecular weight of the PEG chain of the nonionic hydrophilic polymer derivative is preferably 500 to 5,000, more preferably 750 to 3,000. The nonionic hydrophilic polymer chain may be branched or may have a substituent such as a hydroxymethyl group.

[0044] In the lipid particles, the amount of the lipid having a nonionic hydrophilic polymer chain is preferably 0.25 mol % to 12 mol %, more preferably 0.5 mol % to 6 mol %, and even more preferably 1 mol % to 3 mol %, relative to the total amount of lipid.

[0045] The lipid particles may contain nucleic acids. Examples of nucleic acids include plasmid DNA, nanoplasmid DNA, single-stranded DNA, double-stranded DNA, siRNA (small interfering RNA), miRNA (micro RNA), mRNA, antisense oligonucleotides, ribozymes, aptamers, dsRNA, saRNA, sgRNA, shRNA, tRNA, and cyclic RNA, and any of these may be contained. Furthermore, modified nucleic acids may be contained. RNA is particularly preferred as the nucleic acid, and RNA having 5 to 20,000 bases is preferred. In the lipid particles, the mass ratio of lipid to nucleic acid is preferably 2 to 1,000, more preferably 3 to 500, even more preferably 5 to 200, and particularly preferably 5 to 100.

[0046] The method for producing lipid particles is not limited, but can be produced by dissolving all or some of the oil-soluble components of the lipid particle in an organic solvent or the like to form an oil phase, dissolving the water-soluble components in water to form an aqueous phase, and mixing the oil phase and the aqueous phase. A micromixer may be used for mixing, or emulsification may be performed using an emulsifier such as a homogenizer, an ultrasonic emulsifier, a high-pressure injection emulsifier, or the like. Alternatively, lipid particles can be produced by preparing a dried mixture containing lipids by subjecting a lipid-containing solution to vacuum drying using an evaporator or the like or spray drying using a spray dryer, adding this mixture to an aqueous solvent, and further emulsifying using the above-mentioned emulsifier or the like.

[0047] An example of a method for producing lipid particles containing nucleic acid includes the following steps: (a) dissolving the components of lipid particles containing the compound represented by formula (3) or a salt thereof in an organic solvent to obtain an oil phase; (b) mixing the oil phase obtained in step (a) with an aqueous phase containing nucleic acid; (c) diluting the mixture containing the oil phase and aqueous phase obtained in step (b) to obtain a dispersion of nucleic acid-lipid particles; and (d) removing the organic solvent from the dispersion of nucleic acid-lipid particles obtained in step (c).

[0048] Lipid particles refer to particles composed of lipids, and include compositions having any structure selected from lipid aggregates in which lipids are aggregated, micelles, and liposomes, but the structure of the lipid particles is not limited to these as long as the composition contains lipids. Liposomes have a lipid bilayer structure and an aqueous phase inside, and include liposomes with a single bilayer membrane and multi-layered liposomes with multiple layers. Either type of liposome may be included in the present invention.

[0049] The morphology of lipid particles can be confirmed by electron microscope observation or X-ray structural analysis.For example, by using a cryo-transmission electron microscope (cryo-TEM) method, it can be confirmed whether the lipid particles have a lipid bilayer structure (lamellar structure) and an inner water layer, like liposomes, or whether the particles have a core with high electron density inside and a structure packed with lipids and other components.Small-angle X-ray scattering (SAXS) measurement can also be used to confirm whether the lipid particles have a lipid bilayer structure (lamellar structure).

[0050] As an example of the use of lipid particles, nucleic acid (e.g., genes) can be introduced into cells by introducing lipid particles containing nucleic acid into cells. Furthermore, when the lipid particles of the present invention contain nucleic acid having pharmaceutical uses, the lipid particles can be administered to a living body as a nucleic acid drug.

[0051] Lipid particles are highly useful as nucleic acid delivery carriers because they can retain nucleic acids at a high encapsulation rate. According to the nucleic acid delivery carrier of the present invention, for example, the obtained lipid particles can be mixed with nucleic acids and transfected in vitro or in vivo to introduce nucleic acids into cells. The nucleic acid delivery carrier is also useful as a nucleic acid delivery carrier for nucleic acid medicines. That is, the lipid particles are useful as a composition for nucleic acid delivery in vitro or in vivo (preferably in vivo).

[0052] The present invention will now be described with reference to examples, but the present invention is not limited to these examples.

[0053] Unless otherwise specified, purification by column chromatography was performed using an automatic purification system ISOLERA (Biotage), a medium-pressure preparative purification system Purif-espoir-2 (Shoko Science Co., Ltd.), or a medium-pressure liquid chromatograph YFLC W-prep 2XY (Yamazen Corporation). Unless otherwise specified, the carrier used in silica gel column chromatography was Chromatorex Q-Pack SI 50 (Fuji Silysia Chemical Ltd.), or Hi-Flash Column W001, W002, W003, W004, or W005 (Yamazen Corporation). NH silica gel was Chromatorex Q-Pack NH 60 (Fuji Silysia Chemical Ltd.). NMR spectra were measured using a Bruker AVNEO400 (Bruker) using tetramethylsilane as an internal standard, and all δ values ​​are expressed in ppm. MS spectra were measured using an ACQUITY SQD LC / MS System (manufactured by Waters).

[0054] [Example 1] (1)

[0055] To a mixture of 2-pentyl-1-heptanol (15.0 g) and 6-bromohexanoic acid (18.1 g) in toluene (116 mL), p-toluenesulfonic acid monohydrate (1.50 g) was added and heated to reflux for 2 hours. After the reaction mixture was cooled to room temperature, 10% aqueous sodium bicarbonate solution (105 mL) was added. The mixture was then transferred to a separatory funnel and washed with ethyl acetate (105 mL) and water (45 mL). The aqueous layer was discarded and washed with water (105 mL) and saturated brine (45 mL). The organic layer was dried over anhydrous sodium sulfate, and the anhydrous sodium sulfate was removed by filtration. The filtrate was then distilled under reduced pressure at an external temperature of 40°C using an evaporator to obtain oily 2-pentylhexyl-6-bromohexanoic acid (28.7 g). １ H-NMR (CDCl ３) δ: 3.97 (2H, d, J = 5.8Hz), 3.40 (2H, t, J = 6.8Hz), 2.32 (2H, t, J = 7.4Hz), 1.91-1.84 (2H , m), 1.69-1.61 (3H, m), 1.51-1.44 (2H, m), 1.34-1.20 (16H, m), 0.88 (6H, t, J=6.9Hz).

[0056] (2)

[0057] A mixed solution of 2-pentylhexyl-6-bromohexanoic acid (27.0 g) in DMF (20 mL) and ethyl acetate (20 mL) was added dropwise over 20 minutes to a mixture of heptylamine (25.7 g), potassium carbonate (31.4 g), potassium iodide (37.0 g), DMF (80 mL), and ethyl acetate (80 mL), and the mixture was stirred for 2 hours. After adding ethyl acetate (100 mL) and water (160 mL), the mixture was transferred to a separatory funnel, the aqueous layer was discarded, and the organic layer was separated. The mixture was then washed with water (100 mL) and saturated brine (60 mL), dried over anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure. The resulting crude product was purified by silica gel chromatography (hexane-ethyl acetate-methanol) to obtain 2-pentylheptyl-6-(heptylamino)hexanoic acid (23.1 g) as a pale yellow oil. １ H-NMR (CDCl ３ ) δ: 3.96 (2H, d, J = 5.8Hz), 2.61-2.56 (4H, m), 2.31 (2H, t, J = 7.5Hz), 1.68-1.20 (34H, m), 0.88 (9H, m).

[0058] (3)

[0059] A solution of 2-pentylheptyl-6-(heptylamino)hexanoic acid (8.58 g) in tetrahydrofuran (30 mL) was added to a mixture of 1,1'-carbonyldi(1,2,4-triazole) (5.31 g) and tetrahydrofuran (100 mL), and the mixture was stirred at room temperature for 2 hours. Ethyl acetate (150 mL) and water (100 mL) were added, and the organic layer was separated. The resulting organic layer was washed with water (100 mL) and saturated brine (100 mL), then dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting crude product was purified using a column to give 2-pentylheptyl-6-(N-heptyl-1H-1,2,4-triazole-1-carboxamido)hexanoic acid (7.39 g). １ H-NMR (CDCl ３ ) δ: 8.80 (1H, s), 7.98 (1H, s), 3.97 (2H, d, J = 5.8Hz), 3.80-3.25 (4H, m), 2.32 (2H, t, J = 7.4Hz), 1.74-1.17 (33H, m), 0.93-0.82 (9H, m).

[0060] (4)

[0061] To a solution of 2-pentylheptyl-6-(N-heptyl-1H-1,2,4-triazole-1-carboxamido)hexanoic acid (7.39 g) in tetrahydrofuran (50 mL) was added a mixture of 2,2'-((2-(diethylamino)ethyl)azanediyl)bis(ethan-1-ol) (1.40 g) and sodium hydride (1.40 g) in tetrahydrofuran (25 mL) in an ice bath, and the mixture was stirred at room temperature for 30 minutes. A saturated ammonium chloride solution was added in an ice bath, and the mixture was stirred at room temperature for 3 hours. Ethyl acetate (300 mL) and water (200 mL) were added to the reaction mixture, and the organic layer was separated. The organic layer was further washed with water (200 mL) and saturated brine (50 mL). The organic layer was dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The obtained crude product was purified by silica gel chromatography (hexane-ethyl acetate) to obtain oily bis(2-pentylheptyl)-12-(2-(diethylamino)ethyl)-7,17-diheptyl-8,16-dioxo-9,15-dioxa-7,12,17-triazatricosane dioate (4.16 g) in high yield (67%). １H-NMR (CDCl ３ ) δ: 4.10 (4H, t, J = 6.4Hz), 3.96 (4H, d, J = 5.8Hz), 3.25-3.08 (8H, m), 2.79 (4H, t, J = 6.4Hz), 2.68-2.64 (2H, m ), 2.54-2.48 (6H, m), 2.30 (4H, t, J = 7.5Hz), 1.70-1.18 (66H, m), 1.10 (6H, t, J = 7.1Hz), 0.90-0.86 (18H, m). MS m / z (M+H): 1052.

[0062] [Example 2] (1)

[0063] To a solution of triphosgene (69.7 mg) in tetrahydrofuran (4.0 mL), pyridine (57 μL) and 2-pentylheptyl-6-(heptylamino)hexanoic acid (200 mg) were added, and the mixture was stirred at room temperature for 15 minutes. Ethyl acetate (4 mL) and water (2 mL) were added, and the organic layer was separated. The resulting organic layer was washed with water (2 mL) and saturated brine (2 mL), then dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting crude product was purified by silica gel chromatography (hexane-ethyl acetate) to give 2-pentylheptyl-6-((chlorocarbonyl)(heptyl)amino)hexanoic acid (159 mg) as an oil. １ H-NMR (CDCl ３ ) δ: 4.02-3.94 (2H, m), 3.41-3.30 (4H, m), 2.34-2.29 (2H, m), 1.71-1.23 (33H, m), 0.90-0.86 (9H, m).

[0064] (2)

[0065] Using the obtained 2-pentylheptyl-6-((chlorocarbonyl)(heptyl)amino)hexanoic acid as a starting material, bis(2-pentylheptyl)-12-(2-(diethylamino)ethyl)-7,17-diheptyl-8,16-dioxo-9,15-dioxa-7,12,17-triazatricosane dioate was obtained in the same manner as in [Example 1] (4). １ H-NMR (CDCl ３) δ: 4.10 (4H, t, J = 6.4Hz), 3.96 (4H, d, J = 5.8Hz), 3.25-3.08 (8H, m), 2.79 (4H, t, J = 6.4Hz), 2.68-2.64 (2H, m ), 2.54-2.48 (6H, m), 2.30 (4H, t, J = 7.5Hz), 1.70-1.18 (66H, m), 1.10 (6H, t, J = 7.1Hz), 0.90-0.86 (18H, m). MS m / z (M+H): 1052.

[0066] [Example 3] Compounds 1 to 10 were synthesized in the same manner as in Example 1 (1), except that Compound A below was used instead of 6-bromohexanoic acid and Compound B below was used instead of heptylamine in Example 1 (2).

[0067]

[0068] Example 4 Compounds 11 to 20 were synthesized using the amines (see the table below) synthesized in Example 3 as starting materials instead of 2-pentylheptyl-6-(heptylamino)hexanoic acid in Example 1(3).

[0069]

[0070] Example 5 Compounds 21 to 40 were synthesized in the same manner as in Example 1 (4), except that the activated triazole synthesized in Example 4 (see the table below) was used instead of 2-pentylheptyl-6-(N-heptyl-1H-1,2,4-triazole-1-carboxamido)hexanoic acid, and the diol was 2,2'-((2-(diethylamino)ethyl)azanediyl)bis(ethan-1-ol) or 2,2'-((3-(diethylamino)propyl)azanediyl)bis(ethan-1-ol).

[0071]

Claims

1. A compound represented by formula (1). In the formula, R １ represents a hydrogen atom or a hydrocarbon group having 1 to 12 carbon atoms; R ２ represents a divalent hydrocarbon group having 1 to 12 carbon atoms, and R ３ represents a hydrocarbon group having 1 to 24 carbon atoms; X １ teeth, or a halogen atom.

2. R １ represents a hydrocarbon group having 3 to 8 carbon atoms; R ２ represents a divalent hydrocarbon group having 1 to 5 carbon atoms; R ３ The compound according to claim 1, wherein represents a hydrocarbon group having 12 to 24 carbon atoms.

3. X １ The compound according to claim 1, wherein the halogen atom represented by is a chlorine atom.

4. or The compound according to claim 1, which is a compound represented by the formula:

5. A method for producing a compound represented by formula (3) or a salt thereof, comprising reacting a compound represented by formula (1) according to claim 1 with a compound represented by formula (2) or a salt thereof: In the formula, R ４ and R ５ each independently represents a hydrocarbon group having 1 or 2 carbon atoms; R ４ and R ５ each independently represents an optionally substituted hydrocarbon group having 1 to 18 carbon atoms; R ４ and R ５ The substituents on the optionally substituted hydrocarbon group having 1 to 18 carbon atoms represented by the formula (I) are each independently —OH, —COOH, —NR ９ R １０ , -OC(O)OR １１ , -C(O)O-R １２ , —OC(O)—R １３ , -O-R １４ , —C(O)NR １５ R １６ , -NR １７ C(O)R １８ , -N(R １９ ) S (O) ２ R ２０ , -N(R ２１ )C(O)N(R ２２ ) R ２３ , -N(R ２４ ) C(S)N(R ２５ ) R ２６ , -OC(O)N(R ２７ ) R ２８ , or -N(R ２９ )C(O)OR ３０ indicates R ９ and R １０ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R １１ , R １２ , R １３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０ each independently represents a hydrogen atom or an optionally substituted hydrocarbon group having 1 to 24 carbon atoms; R １１ , R １２ , R １３ , R １４ , R １５ , R １６ , R １７ , R １８ , R １９ , R ２０ , R ２１ , R ２２ , R ２３ , R ２４ , R ２５ , R ２６ , R ２７ , R ２８ , R ２９ , and R ３０ The substituent on the optionally substituted hydrocarbon group having 1 to 24 carbon atoms represented by is an aryl group having 6 to 20 carbon atoms, a heterocyclic group, —OH, —COOH, or —NR ３１ R ３２ indicates R ３１ and R ３２ each independently represents a hydrogen atom or a hydrocarbon group having 1 to 8 carbon atoms; R ４ and R ５ , or R ４ and R ６ may be joined together to form a 4- to 7-membered ring, R ６ represents a hydrocarbon group having 2 to 4 carbon atoms, and R ７ and R ８ each independently represents a hydrocarbon group having 2 or 3 carbon atoms, or R ５ and R ６ may be linked to form a cyclic structure, In the formula, R １ , R ２ and R ３ has the same meaning as defined in claim 1, and R ４ , R ５ , R ６ , R ７ and R ８ is defined as in formula (2).

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