Piperazine compounds

Abstract

The present invention particularly relates to novel piperazine compounds of formula (I) or (II). These piperazine compounds can be used, for example, in lipid nanoparticle compositions for drug delivery and cancer treatment. TIFF2025531749000244.tif90149

Description

[Technical Field]

[0001] This application describes novel piperazine compounds that can be used, for example, in lipid nanoparticle compositions for drug delivery and cancer treatment.

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 63 / 375,061, filed September 9, 2022, the disclosure of which is incorporated herein by reference in its entirety. [Background technology]

[0003] For therapeutics, diagnostics, reagents, and biological assays, it is of great interest to be able to deliver nucleic acids into cells, for example, to trigger intracellular translation of the nucleic acid and production of the encoded protein. Nucleic acid delivery has been widely explored as a potential therapeutic option for certain pathologies. In particular, double-stranded DNA (dsDNA) therapy has become an increasingly important option for the treatment of various diseases, including cancer. Intracellular delivery of dsDNA can stimulate the innate immune response, which may be useful in the treatment of various diseases. Summary of the Invention [Problem to be solved by the invention]

[0004] Therefore, there is a need to develop lipid compounds that can be useful for facilitating the delivery of nucleic acids to target cells or subjects in need thereof. The present invention fulfills this unmet need. [Means for solving the problem]

[0005] In a general aspect, the present application relates to a compound of formula (I) [ka] or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, n is 0 or 1, X1 and X2 are different or identical and each independently represent CHR3; R1 and R2 are different or identical, and C 1~8 Alkyl and -C 1~8 alkyl-Y, where Y is -OR, -SR, or N(R) 2 and the above C 1~8 Alkyl is halogen and C 1~8 optionally substituted with one or more selected from the group consisting of alkyl; R3, independently at each occurrence, is selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, -SH, alkoxyl, alkylthio, aryl, cycloalkyl, and heterocycle, wherein said aryl, cycloalkyl, or heterocycle is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, alkoxy, and amino; R4, independently in each occurrence, is selected from alkyl and -C 1~4 alkyl-Z, wherein Z is -OR, -SR, or N(R) 2 wherein the alkyl is optionally substituted with one or more hydroxy groups; R5, in each occurrence, is independently selected from hydroxyl-substituted alkyl and (—C 1~4 Alkyl-O) m -C 1~4 alkyl-N(R6)2, m is 0 or 1, and R6 is hydroxyl-substituted alkyl; However, at least one of R1 and R2 is C 1~4 not alkyl].

[0006] In some embodiments, X1 and X2 are the same, preferably CH2.

[0007] In some embodiments, the compound of formula (I) is a compound of formula (IA) [ka] [wherein R1, R2, and n are defined as above].

[0008] In some embodiments, n is 0.

[0009] In some embodiments, R is -C 1~4 It is alkyl-Y.

[0010] In some embodiments, R2 is -C 1~4 It is alkyl-Y.

[0011] In some embodiments, Y is -OR4, -SR4, or N(R4)2, preferably SR4.

[0012] In some embodiments, R4 is -C 1~4 It is alkyl-Z.

[0013] In some embodiments, Z is N(R5)2.

[0014] In some embodiments, R5 is hydroxyl-substituted alkyl.

[0015] In some embodiments, R3 at each occurrence is independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, -SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

[0016] In some embodiments, n is 0 and the compound is a compound of formula (I-A1). [ka]

[0017] In other embodiments, n is 1 and the compound is a compound of formula (I-A2). [ka]

[0018] In some embodiments, the compound of formula (I) is a compound of formula (IB) [ka] [Wherein R3 and R 3’ are each independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy, and R and R are as defined above.

[0019] In certain embodiments, the compound of formula (I) is a compound of formula (I-B1) [ka] [Wherein R3 and R 3’are different or the same and are each independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

[0020] In a further embodiment, R and R 3’ are identical.

[0021] In another general aspect, the application provides a compound of formula (II) [ka] or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, p is an integer from 1 to 10, q is an integer from 3 to 8; R8 is unbranched or branched alkyl.

[0022] In another aspect, the present application relates to a pharmaceutical composition comprising a compound described herein or a stereoisomer, tautomer, pharmaceutically acceptable salt, or solvate thereof, and a pharmaceutically acceptable carrier.

[0023] In another aspect, the application relates to the use of the compounds described herein, or stereoisomers, tautomers, pharmaceutically acceptable salts, or solvates thereof, for drug delivery or cancer treatment.

[0024] Other features and advantages of the present invention will be apparent from the further description provided herein, including the different examples. The examples provided illustrate different components and methodologies useful in carrying out the invention. Such examples do not limit the claimed invention. Based on this disclosure, one of ordinary skill in the art will be able to identify and use other components and methodologies useful in carrying out the invention.

[0025] The foregoing summary, as well as the following detailed description of preferred embodiments of the present application, will be better understood when read in conjunction with the accompanying drawings. It should be understood, however, that the present application is not limited to the precise embodiments shown in the drawings. [Brief explanation of the drawings]

[0026] [Figure 1] 1 shows plasma hEPO levels assessed 4 hours after administration of LNP-dsDNA formulations LNP1 to 11 using the SIL1 series of lipids. [Figure 2] 1 shows hFIX levels in HepG2 cells incubated with 75 ng / well of LNP-dsDNA formulations LNP915-934 using SIL2 series lipids. [Figure 3] 1 shows hFIX levels in HepG2 cells incubated with 150 ng / well of LNP-dsDNA formulation LNP915-934 using the SIL2 series of lipids. [Figure 4] IRF levels of THP-1 cells incubated with 150 ng / well of LNP-dsDNA formulation LNP915-934 using SIL2 series lipids are shown. [Figure 5] IRF levels of THP-1 cells incubated with 75 ng / well of LNP-dsDNA formulations LNP915-934 using SIL2 series lipids are shown. [Figure 6] NFkB levels in THP-1 cells incubated with 150 ng / well of LNP-dsDNA formulations LNP915-934 using SIL2 series lipids are shown. [Figure 7] NFkB levels in THP-1 cells incubated with 75 ng / well of LNP-dsDNA formulations LNP915-934 using SIL2 series lipids are shown. [Figure 8] 1 shows plasma hEPO levels assessed 4 hours after administration of LNP-mRNA formulations LNP31 to 42 using the SIL3 series lipids. [Figure 9] 1 shows plasma hEPO levels assessed 1 week and 2 weeks after administration of LNP-DNA formulations LNP43-54 using SIL3 series lipids. DETAILED DESCRIPTION OF THE INVENTION

[0027] Various publications, articles, and patents are cited or described in the Background and throughout the specification, and each of these references is incorporated herein by reference in its entirety. The discussion of documents, acts, materials, devices, articles, and the like which has been included herein is for the purpose of providing a context for the present disclosure. Such discussion is not an admission that any or all of these items constitute part of the prior art with respect to the present disclosure.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Unless otherwise defined, certain terms used herein have the meanings set forth herein. All patents, published patent applications, and publications cited herein are incorporated by reference as if fully set forth herein.

[0029] It must be noted that as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise.

[0030] Unless otherwise specified, the term "at least" following a series of elements should be understood to refer to every element in the series. For example, the phrase "at least A, B, and C" means that each of A, B, and C is present. The term "at least one of" following a series of elements should be understood to refer to either a single element in the series or any combination of two or more elements in the series. For example, the phrase "at least one of A, B, and C" means that only A is present, only B is present, only C is present, both A and B are present, both A and C are present, both B and C are present, or each of A, B, and C is present. Depending on the context, "at least one of" following a series of elements can also encompass situations where any one or more of the elements are present in more than one instance; for example, "at least one of A, B, and C" can also encompass situations where A is present multiple times alone or in further combination with any one or more of the elements B and C.

[0031] As used herein, the conjunction "and / or" between multiple listed elements is understood to encompass both individual and combined options. For example, when two elements are joined by "and / or," the first option indicates that the first element is applicable without the second element. The second option indicates that the second element is applicable without the first element. The third option indicates that the first and second elements are applicable together. Any one of these options is understood to be within the meaning and therefore meets the requirements of the term "and / or" as used herein. Two or more of the options may also be simultaneously applicable and therefore meet the requirements of the term "and / or."

[0032] Unless otherwise stated, any numerical values, such as concentrations or concentration ranges described herein, should be understood in all cases as being modified by the term "about." Thus, numerical values ​​typically include ±10% of the described value. For example, the description "10 times" includes 9 times and 11 times. As used herein, the use of numerical ranges includes all possible subranges, including integers and fractions of values ​​within such ranges, and all individual numerical values ​​within the range, unless otherwise clearly indicated by the context.

[0033] As used herein, "subject" means any animal, e.g., a mammal, that will be treated or has been treated by the methods described herein. The term "mammal," as used herein, encompasses any mammal. Examples of mammals include, but are not limited to, cows, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, and non-human primates (NHPs), such as monkeys or apes, humans, etc.

[0034] The phrase "pharmaceutically acceptable salt" refers to a salt of a subject compound that is safe and effective for topical administration to mammals and that possesses the desired biological activity. Pharmaceutically acceptable salts include salts of acidic or basic groups present in the specified compound. Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, superphosphate, isonicotinate, carbonate, bicarbonate, acetate, lactate, salicylate, citrate, tartrate, propionate, butyrate, pyruvate, oxalate, malonate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate (i.e., 1,1'-methylene-bis-(2-hydroxy-3-naphthoate)) salts. Certain compounds used in this application can form pharmaceutically acceptable salts with various amino acids. Suitable base salts include, but are not limited to, aluminum, calcium, lithium, magnesium, potassium, sodium, zinc, bismuth, and diethanolamine salts. For a review of pharmaceutically acceptable salts, see Berge et al., 66 J.Pharm.Sci.1-19 (1977), which is incorporated herein by reference.

[0035] As used herein, the term "alkyl" means a saturated, monovalent, unbranched or branched hydrocarbon chain. An alkyl group can be unsubstituted or substituted with one or more suitable substituents. Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl, isopropyl), butyl (e.g., n-butyl, isobutyl, tert-butyl), and pentyl (e.g., n-pentyl, isopentyl, neopentyl), and the like. An alkyl group can have a specified number of carbon atoms. When a number appears in a subscript after the symbol "C," the subscript further defines the number of carbon atoms that particular alkyl can contain. For example, "C1-C 10 Alkyl" or "C 1~10 "Alkyl" is intended to include alkyl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms. In addition, "C1-C8 alkyl" or "C 1~8 "Alkyl" refers to an alkyl having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms.

[0036] The term "cycloalkyl" refers to any stable monocyclic or polycyclic saturated hydrocarbon ring system. A cycloalkyl group can be unsubstituted or substituted with one or more suitable substituents. A cycloalkyl group can have a specified number of carbon atoms. For example, "C3-C6 cycloalkyl" or "C 3~6"Cycloalkyl" includes cycloalkyl groups having 3, 4, 5, or 6 ring carbon atoms, i.e., cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. Polycyclic cycloalkyl includes bridged, fused, and spiro ring structures in which all ring atoms are carbon atoms. A "spirocycle" is a polycyclic ring system in which two rings share one carbon atom (called a "spiroatom," typically a quaternary carbon atom). A "fused ring" is a polycyclic ring system in which two rings share two adjacent atoms (called "bridgehead atoms"), i.e., the two rings share one covalent bond such that the bridgehead atoms are directly connected. A "bridgehead ring" is a polycyclic ring system in which two rings share three or more atoms, with the bridgehead atoms separated by a bridge containing at least one atom. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like.

[0037] As used herein, the term "alkenyl" means a hydrocarbon straight or branched chain containing at least one carbon-carbon double bond and, optionally, having a specified number of carbon atoms (i.e., C2-C4 alkenyl or C 2~4 Alkenyl means an alkenyl having 2 to 4 carbon atoms).

[0038] As used herein, the term "alkynyl" means a hydrocarbon straight or branched chain containing at least one carbon-carbon triple bond and, optionally, having a specified number of carbon atoms (i.e., C2-C4 alkenyl or C 2~4 Alkenyl means an alkenyl having 2 to 4 carbon atoms).

[0039] The term "aryl," as used herein, refers to a group containing any carbon-based aromatic group, including, but not limited to, phenyl, naphthyl, anthracenyl, phenanthranyl, and the like. Aryl moieties are well known and are described, for example, in Lewis, RJ (ed.), Hawley's Condensed Chemical Dictionary, 13th Edition, John Wiley & Sons, Inc., New York (1997). Aryl groups can be substituted or unsubstituted with one or more suitable substituents. Aryl groups can include a single ring structure (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic or tricyclic). For example, an aryl group can be a monocyclic aryl group, such as phenyl.

[0040] The term "heterocyclyl" includes stable monocyclic and polycyclic hydrocarbons containing at least one heteroatom ring member, such as sulfur, oxygen, or nitrogen; the ring structures are saturated or partially unsaturated, although the ring systems are not necessarily aromatic. Heterocyclyl groups can be unsubstituted or substituted with one or more suitable substituents at any one or more of the carbon atoms and / or nitrogen heteroatoms of the heterocyclyl. Heterocyclyls can include single ring structures (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic). Polycyclic heterocyclyls include bridged, fused, and spiro ring structures in which at least one ring atom of at least one of the rings of the polycyclic ring system is a heteroatom, such as oxygen, nitrogen, or sulfur; bridged, fused, and spiro rings are as defined above. The heterocyclyl ring can be attached to the parent molecule at any suitable heteroatom (typically nitrogen) or carbon atom of the ring. The term "4-9 membered monocyclic or bicyclic heterocyclyl" includes any 4-, 5-, 6-, 7-, 8-, or 9-membered monocyclic or bicyclic ring structure containing at least one heteroatom ring member selected from oxygen, nitrogen, and sulfur, or independently selected from oxygen and nitrogen, and optionally containing 1 to 3 additional heteroatoms independently selected from oxygen, nitrogen, and sulfur, or independently selected from oxygen and nitrogen, wherein the ring structure is saturated or partially unsaturated, although the ring structure is not necessarily aromatic.

[0041] As used herein, the term "heteroaryl" includes stable monocyclic and polycyclic aromatic hydrocarbons containing at least one heteroatom ring member, such as sulfur, oxygen, or nitrogen. Heteroaryl groups can be unsubstituted or substituted with one or more suitable substituents. Heteroaryl can include a single ring structure (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic or tricyclic). Each ring of a heteroatom-containing heteroaryl group can contain one or two oxygen or sulfur atoms and / or one to four nitrogen atoms, provided that the total number of heteroatoms in each ring is four or less and each ring has at least one carbon atom. Heteroaryl groups that are polycyclic, e.g., bicyclic or tricyclic, must contain at least one fully aromatic ring, but the other fused ring(s) may or may not be aromatic. For example, for a bicyclic heteroaryl, the fused rings completing the bicyclic group can contain only carbon atoms and may be saturated, partially saturated, or unsaturated. A heteroaryl can be attached to the parent molecule at any available nitrogen or carbon atom of any ring of the heteroaryl group. In some embodiments, the term "heteroaryl" refers to 5- or 6-membered monocyclic groups and 9- or 10-membered bicyclic groups having at least one heteroatom (O, S, or N) in at least one of the rings, where the heteroatom-containing ring typically has 1, 2, or 3 heteroatoms selected from O, S, and / or N, e.g., 1 or 2 heteroatoms. A heteroaryl group can be unsubstituted or substituted with one or more suitable substituents on any one or more of the carbon and / or nitrogen heteroatoms of the heteroaryl. The nitrogen and sulfur heteroatoms of a heteroaryl can be optionally oxidized (i.e., N→O and S(O)r, where r is 0, 1, or 2).

[0042] The term "alkoxy," as used herein, refers to an -O-alkyl group, where alkyl is as defined above. An alkoxy group is attached to the parent molecule via a bond to an oxygen atom. An alkoxy group can have a specified number of carbon atoms. For example, "C1-C 10 Alkoxy" or "C 1~10 "Alkoxy" is intended to include alkoxy groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms. Additionally, examples of "C1-C4 alkoxy" or "C 1~4 "Alkoxy" refers to an alkoxy having 1, 2, 3, or 4 carbon atoms. Examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy (e.g., n-propoxy, isopropoxy), butoxy (e.g., n-butoxy, isobutoxy, tert-butoxy), pentyloxy (e.g., n-pentyloxy, isopentyloxy, neopentyloxy), and the like. An alkoxy group can be unsubstituted or substituted with one or more suitable substituents. Similarly, "alkylthio" or "thioalkoxy" refers to an alkyl group, as defined above, attached to the parent molecule via a bond to a sulfur atom, such as -S-methyl, -S-ethyl, and the like. Representative examples of alkylthio include, but are not limited to, -SCH, -SCHCH, and the like.

[0043] As used herein, the term "halogen" means fluorine, chlorine, bromine, or iodine. Correspondingly, the term "halo" means fluoro, chloro, bromo, and iodo.

[0044] "Haloalkyl" is intended to include both branched and straight-chain saturated aliphatic hydrocarbon radicals substituted with one or more halogen atoms. "Fluorinated alkyl" or "fluoroalkyl" specifically refers to any alkyl group as defined above substituted with at least one fluoro atom, for example, 1 to 3 fluoro atoms, for example, 1, 2, or 3 fluoro atoms. Examples of haloalkyl include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, pentachloroethyl, 2,2,2-trifluoroethyl, heptafluoropropyl, and heptachloropropyl. Suitable examples of fluoroalkyl include, but are not limited to, -CF3, -CHF2, -CH2F, -CH2CF3, -CF2CF3, and the like.

[0045] The terms "hydroxy" and "hydroxyl" can be used interchangeably and refer to --OH.

[0046] The term "carboxy" refers to --COOH.

[0047] The term "ester" refers to -COOR, where R is alkyl as defined above.

[0048] The term "cyano" refers to -CN.

[0049] The term "oxo" refers to a double-bonded oxygen group, ie, a substituent of formula =O.

[0050] The term "keto" refers to -C(O)R, where R is alkyl as defined above.

[0051] The term "amino" refers to -NH2. One or more hydrogen atoms of the amino group may be replaced by a substituent such as an alkyl group, which is called an "alkylamino." An alkylamino group has one or both hydrogen atoms of the amino group replaced with an alkyl group and is attached to the parent molecule via a bond to the nitrogen atom of the alkylamino group. For example, alkylamino includes methylamino (-NHCH3), dimethylamino (-N(CH3)2), -NHCH2CH3, and the like.

[0052] The term "aminoalkyl," as used herein, is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups substituted with one or more amino groups. For example, "C 1~4 "Aminoalkyl" is intended to include alkyl groups having 1, 2, 3, or 4 carbon atoms substituted with one or more amino groups. The aminoalkyl group is attached to the parent molecule via a bond to a carbon atom of the alkyl portion of the aminoalkyl group. Representative examples of aminoalkyl groups include, but are not limited to, -CHNH, -CHCHNH, and -CHCH(NH)CH.

[0053] As used herein, "amide" refers to -C(O)N(R)2, where each R is independently an alkyl group (including both branched and straight chain alkyl groups) or a hydrogen atom. Examples of amide groups include, but are not limited to, -C(O)NH2, -C(O)NHCH3, and -C(O)N(CH3)2.

[0054] The terms "hydroxyl-substituted alkyl," "hydroxylalkyl," and "hydroxyalkyl" are used interchangeably and refer to branched or straight-chain aliphatic hydrocarbon groups substituted with one or more hydroxy groups. The hydroxyalkyl group is attached to the parent molecule via a bond to a carbon atom of the alkyl portion of the hydroxyalkyl group. The hydroxyalkyl group can have a specified number of carbon atoms. For example, "C1-C 10 Hydroxyalkyl" or "C 1~10"Hydroxyalkyl" is intended to include hydroxyalkyl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms. In addition, "C1-C4 hydroxylalkyl" or "C 1~4 "Hydroxyalkyl" refers to a hydroxyalkyl group having 1, 2, 3, or 4 carbon atoms. Examples of hydroxyalkyl include, but are not limited to, hydroxymethyl (-CHOH), hydroxyethyl (-CHCHOH), and the like.

[0055] According to the conventions used in the art, [ka] is used in structural formulas herein to depict a bond that is the point of attachment of a group, moiety, or substituent to the core, backbone, or parent molecular structure.

[0056] When a bond to a substituent is shown to cross the bond connecting two atoms in a ring, then such substituent may be bonded to any atom on the ring.

[0057] The term "substituted," as used herein with respect to any organic radical (e.g., alkyl, cycloalkyl, heteroaryl, aryl, heterocyclyl, etc.), means that at least one hydrogen atom is replaced with a non-hydrogen group, provided that all normal valences are maintained and the substitution results in a stable compound. When a particular group is "substituted," the group can have one or more substituents, such as 1 to 5 substituents, 1 to 3 substituents, or 1 to 2 substituents, independently selected from a list of substituents. The term "independently," when used in reference to substituents, means that when two or more such substituents may be present, such substituents may be the same or different from one another. Examples of suitable substituents include, but are not limited to, alkyl, halo, haloalkyl, alkoxy, amido, hydroxy, hydroxyalkyl, amino, carboxyl, ester, oxo, cyano, and the like.

[0058] When any variable occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0 to 3 R groups, then that group may be optionally substituted with up to 3 R groups, and R is independently selected from the definitions of R at each occurrence.

[0059] The term "optional" or "optionally" means that the subsequently described event or circumstance may, but need not, occur, and such a description includes situations in which the event or circumstance occurs or does not occur. For example, "optionally substituted heterocyclyl" means that a substituent may, but need not, be present, and such a description includes situations in which the heterocyclyl group is substituted with a suitable substituent and situations in which the heterocyclyl group is not substituted with any substituent.

[0060] Those skilled in the art will recognize that in certain embodiments, the compounds described herein can have one or more asymmetric carbon atoms within their structure. As used herein, any chemical formula with bonds shown as solid lines only, and not as solid wedges or hashed wedges, or otherwise shown as having a particular configuration (e.g., R or S) around one or more atoms, contemplates each possible stereoisomer or a mixture of two or more stereoisomers. Stereoisomers include enantiomers and diastereomers. Enantiomers are stereoisomers that are non-superimposable mirror images of each other. A 1:1 mixture of a pair of enantiomers is a racemate or racemic mixture. Diastereomers (or diastereoisomers) are stereoisomers that are not enantiomers, i.e., they are not related as mirror images, and occur when two or more stereoisomers of a compound have different configurations at one or more equivalent stereocenters and are not mirror images of each other. Substituents (eg, alkyl, heterocyclyl, etc.) can contain stereocenters in either the R or S configuration.

[0061] Certain examples include chemical structures that include the terms (R) or (S). When (R) or (S) is used in the name of a compound or in the chemical designation of a compound, it is intended to mean that the compound is a single isomer at that stereocenter, and that either the (R) or (S) absolute configuration has been established.

[0062] Stereochemically pure isomeric forms can be obtained by techniques known in the art in light of the present disclosure. For example, diastereomeric isomers can be separated by physical separation methods, such as fractional crystallization and chromatographic techniques, and enantiomers can be separated from each other by selective crystallization of diastereomeric salts with optically active acids or bases, or by chiral chromatography. Pure stereoisomers can also be synthetically prepared from appropriate stereochemically pure starting materials or by using stereoselective reactions.

[0063] The compounds described herein may also form tautomers. The term "tautomer" refers to interchangeable forms of a particular compound structure, differing in the displacement of hydrogen atoms and electrons. Tautomers are constitutional isomers of chemical compounds that readily interconvert, usually resulting in the rearrangement of protons (hydrogen). Thus, two structures can be in equilibrium through the shifting of π electrons and atoms (usually hydrogen). All tautomers and mixtures of tautomers of the compounds described herein are within the scope of this application.

[0064] The compounds described herein can exist in solvated and unsolvated forms. The term "solvate" refers to a physical association of a compound of the present application with one or more solvent molecules, for example, through hydrogen bonding. The solvent molecules in a solvate can be in an ordered and / or disordered arrangement. A solvate can contain either stoichiometric or non-stoichiometric amounts of solvent molecules. "Solvate" encompasses both solution-phase and isolatable solvates. The compounds of the present application can form solvates with water (i.e., hydrates) or common organic solvents. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates. Solvation methods are generally known in the art.

[0065] Additionally, all isotopes of atoms occurring in the compounds described herein are included within the scope of this application. Isotopes include atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include deuterium and tritium. Isotopes of carbon include 13 C and 14C. Isotopically labeled compounds can generally be prepared by conventional techniques known to those skilled in the art, or by processes analogous to those described herein, using an appropriate isotopically labeled reagent in place of the otherwise unlabeled reagent.

[0066] As used herein, compound names are intended to encompass all possible isomeric forms, including stereoisomers of the compound (e.g., enantiomers, diastereomers, racemates or racemic mixtures, and mixtures thereof).

[0067] compound In one general aspect, the present application provides a compound of formula (I) [ka] or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, n is 0 or 1, X1 and X2 are different or identical and each independently represent CHR3; R1 and R2 are different or identical, and C 1~4 Alkyl and -C 1~4 alkyl-Y, where Y is -OR, -SR, or N(R) 2 and the above C 1~4 Alkyl is halogen and C 1~4 optionally substituted with one or more selected from the group consisting of alkyl; R3, independently at each occurrence, is selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, -SH, alkoxyl, alkylthio, aryl, cycloalkyl, and heterocycle, wherein said aryl, cycloalkyl, or heterocycle is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, alkoxy, and amino; R4, independently in each occurrence, is selected from alkyl and -C 1~4 alkyl-Z, wherein Z is -OR, -SR, or N(R) 2 wherein the alkyl is optionally substituted with one or more hydroxy groups; R5, in each occurrence, is independently selected from hydroxyl-substituted alkyl and (—C 1~4 Alkyl-O) m -C 1~4 alkyl-N(R6)2, m is 0 or 1, and R6 is hydroxyl-substituted alkyl; However, at least one of R1 and R2 is C 1~4 not alkyl].

[0068] In some embodiments, X1 and X2 are the same. In certain embodiments, X1 and X2 are both CH2.

[0069] In some embodiments, the compound of formula (I) is a compound of formula (IA) [ka] [wherein R1, R2, and n are defined as above].

[0070] In some embodiments, n is 0 and the compound is a compound of formula (I-A1). [ka]

[0071] In other embodiments, n is 1 and the compound is a compound of formula (I-A2). [ka]

[0072] In some embodiments, R is C 1~4 Alkyl, for example, methyl, ethyl, propyl (eg, n-propyl, isopropyl), and butyl (eg, n-butyl, isobutyl, tert-butyl).

[0073] In some embodiments, R1 is -C 1~4 alkyl-Y, where Y is -OR, -SR, or N(R) 2 and C 1~4 Alkyl is halogen and C 1~4 and optionally substituted with one or more selected from the group consisting of alkyl.

[0074] In some embodiments, R2 is C 1~4 Alkyl, for example, methyl, ethyl, propyl (eg, n-propyl, isopropyl), and butyl (eg, n-butyl, isobutyl, tert-butyl).

[0075] In some embodiments, R2 is -C 1~4 alkyl-Y, where Y is -OR, -SR, or N(R) 2 and C 1~4 Alkyl is halogen and C 1~4 and optionally substituted with one or more selected from the group consisting of alkyl.

[0076] In some embodiments, Y is -OR4, -SR4, or N(R4)2, preferably SR4.

[0077] In some embodiments, R4 is alkyl, wherein the alkyl is optionally substituted with one or more hydroxy groups.

[0078] In some embodiments, R4 is -C 1~4 alkyl-Z, where Z is -OR, -SR, or N(R) 2 is.

[0079] In certain embodiments, when Y is N(R), R is alkyl, which is optionally substituted with one or more hydroxy groups. In preferred embodiments, R is: [ka] and R7 is unbranched alkyl. In a more preferred embodiment, R4 is [ka] is.

[0080] In certain embodiments, when Y is -N(R4)2, R4 is -C 1~4 alkyl-Z, where Z is -OR, -SR, or N(R) 2 is.

[0081] In certain embodiments, when Y is -OR4 or -SR4, R4 is -C 1~4 alkyl-Z, where Z is -OR, -SR, or N(R) 2 is.

[0082] In some embodiments, R5 is hydroxyl-substituted alkyl.

[0083] In some embodiments, R5 is (-C 1-4 Alkyl-O) m -C 1-4 alkyl-N(R6)2, m is 0 or 1, and R6 is a hydroxyl-substituted alkyl.

[0084] In certain embodiments, Z is N(R5). 2In a preferred embodiment, R5 is: [ka] and R7 is unbranched alkyl. In a more preferred embodiment, R5 is [ka] is.

[0085] In certain embodiments, Z is N(R5). 2 If so, R5 is (-C 1-4 Alkyl-O) m -C 1-4 alkyl-N(R6)2, m is 0 or 1, and R6 is a hydroxyl-substituted alkyl.

[0086] In certain embodiments, when Z is -OR5 or -SR5, R5 is (-C 1~4 Alkyl-O) m -C 1~4 alkyl-N(R6)2, m is 0 or 1, and R6 is a hydroxyl-substituted alkyl.

[0087] In a preferred embodiment, R6 is [ka] and R7 is unbranched alkyl. In a more preferred embodiment, R6 is [ka] is.

[0088] Exemplary compounds of formula (IA) include, but are not limited to, the following compounds SIL1-1 to SIL1-17, and any tautomers, stereoisomers, pharmaceutically acceptable salts, or solvates thereof: [ka] TIFF2025531749000021.tif150149 TIFF2025531749000022.tif125149 TIFF2025531749000023.tif103149 TIFF2025531749000024.tif130149

[0089] In some embodiments, X and X are CHR, and R is independently in each occurrence selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein the alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, -SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein the aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

[0090] In some embodiments, the compound of formula (I) is a compound of formula (IB) [ka] [Wherein R3 and R 3’are each independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy, and R and R are as defined above.

[0091] In certain embodiments, R and R 3’ are identical.

[0092] In certain embodiments, R and R are the same and preferably are -C 1~4 It is alkyl-Y.

[0093] In a further embodiment, Y is -OR4, -SR4, or N(R4)2, preferably SR4.

[0094] In a further embodiment, R4 is -C 1~4 It is alkyl-Z.

[0095] In a further embodiment, Z is N(R5)2.

[0096] In a further embodiment, R5 is hydroxyl-substituted alkyl. In a preferred embodiment, R5 is [ka] and R7 is unbranched alkyl. In a more preferred embodiment, R5 is [ka] is.

[0097] In some embodiments, the compound of formula (IB) is a compound of formula (I-B1) [ka] [Wherein R3 and R 3’ are different or the same and are each independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

[0098] In some embodiments, in the compound of Formula (I-B1), R3 and R3 are the same.

[0099] In some embodiments, the compound of formula (IB) is a compound of formula (I-B2) [ka] wherein R3 is selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, -SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

[0100] Exemplary compounds of formula (IB) include, but are not limited to, the following compounds SIL2-1 to SIL2-22, and any tautomers, stereoisomers, pharmaceutically acceptable salts, or solvates thereof: [ka] TIFF2025531749000031.tif201149 TIFF2025531749000032.tif197149 TIFF2025531749000033.tif201149 TIFF2025531749000034.tif193149 TIFF2025531749000035.tif192149 TIFF2025531749000036.tif203149 TIFF2025531749000037.tif67149

[0101] In another general aspect, the application provides a compound of formula (II) [ka] or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, p is an integer from 1 to 10, q is an integer from 3 to 8; R8 is unbranched or branched alkyl.

[0102] In some embodiments, q is 1-5, preferably 3.

[0103] In some embodiments, p is an integer from 1 to 3, for example, 1, 2, or 3.

[0104] In some embodiments, R is unbranched alkyl, preferably C 2~12 It is an unbranched alkyl.

[0105] In some embodiments, R is branched alkyl, preferably C 2~12 It is a branched alkyl.

[0106] Exemplary compounds of formula (II) include, but are not limited to, the following compounds SIL3-1 to SIL3-11, and any tautomers, stereoisomers, pharmaceutically acceptable salts, or solvates thereof: [ka] TIFF2025531749000040.tif82149

[0107] All possible combinations of the above embodiments of the compounds of Formula (I) or Formula (II), and their tautomers, stereoisomers, pharmaceutically acceptable salts and solvates, are considered to be within the scope of this application.

[0108] Preparation method The compounds described herein can be prepared by any number of processes, generally described below and illustrated in more detail by exemplary compounds in the Examples section of the present specification. The compounds provided herein, as prepared by the processes described below, can be synthesized in the form of mixtures of stereoisomers (e.g., enantiomers, diastereomers), including racemic mixtures of enantiomers, which can be separated from one another using resolution procedures known in the art, including, for example, liquid chromatography using a chiral stationary phase. Additionally or alternatively, stereochemically pure isomeric forms of the compounds described herein can be derived from the corresponding stereochemically pure isomeric forms of the appropriate starting materials, intermediates, or reagents. For example, if a specific stereoisomer is desired, that compound can typically be synthesized by stereospecific preparative methods using stereochemically pure starting materials or intermediate compounds.

[0109] Pharmaceutically acceptable salts of the compounds of the present application can be synthesized from the parent compound that contains an acidic or basic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of the appropriate acid or base in water or an organic solvent, or a mixture of the two. Examples of suitable organic solvents include, but are not limited to, ether, ethyl acetate (EtOAc), ethanol, isopropanol, or acetonitrile.

[0110] By way of example, and not limitation, compounds of Formula (I) or Formula (II) described herein can be prepared according to the following general preparative procedures, as shown in Schemes 1-3, and the examples set forth in this application. To obtain the various compounds of Formula (I) or Formula (II) described herein, one skilled in the art will recognize that starting materials can be appropriately selected to provide the desired product, such that the ultimately desired substituents are carried through the reaction scheme (i.e., are stable throughout the course of the synthesis), with or without protection as appropriate. Alternatively, it may be necessary or desirable to use, in place of the ultimately desired substituent, a suitable group that is carried through the reaction scheme (i.e., is stable throughout the course of the synthesis) and can be replaced with the desired substituent as needed.

[0111] Unless otherwise specified, the variables in Schemes 1-3 are as defined above for the various embodiments of compounds of Formula (I) or Formula (II). If no temperature or temperature range is stated, it should be understood that the reaction is carried out at room temperature.

[0112] [ka] As shown in Scheme 1, a compound of formula (I-B2-2) can be prepared from a compound of formula (I-B2-1) by reaction with compound 1a under suitable reaction conditions, where R3 is defined as above. Compound (I-B2-2) can then be deprotected under suitable conditions, such as with TFA in dichloromethane solvent, to obtain a compound of formula (I-B2-3). A ring-forming reaction of compound (I-B2-3) can yield cyclic compound (I-B2-4) under suitable reaction conditions, such as with pyridine in dichloromethane. Compound (I-B2-4) can be reduced with LAH, followed by a substitution reaction with compound 3a to produce compound I-B2-6. Conversion of compound (I-B2-6) to a compound of formula (I-B2) can be achieved by deprotection followed by further substitution with a compound of formula 2a under suitable reaction conditions.

[0113] [ka] Alternatively, the compound of formula (II-B) can be prepared as shown in Scheme 2. The compound of formula (I-B2-2') is prepared from the compound of formula (I-B2-1) by reaction with compound B under suitable reaction conditions, where R3 is defined as above. The compound of formula (I-B2-2') is then deprotected under suitable conditions, such as with TFA in dichloromethane solvent, to obtain the compound of formula (I-B2-3'). The ring-forming reaction of compound (I-B2-3') gives the cyclic compound (I-B2-4) under suitable reaction conditions, such as with pyridine in dichloromethane. The conversion of the compound of formula (I-B2-4) to the compound of formula (I-B2) can then be similarly achieved via a synthetic route, as shown in Scheme 2.

[0114] [ka] As shown in Scheme 3, compounds of formula (II-3) are prepared from compounds of formula (II-1) by reaction with alcohol II-2 under suitable ester bond forming conditions, where R is defined as above. Compounds of formula (II-3) are then epoxidized by reaction with m-CPBA in a suitable solvent, such as dichloromethane, to give compounds of formula (II-4). Conversion of compounds of formula (II-4) to compounds of formula (II) is achieved by reaction with compounds of formula (II-5) in a suitable solvent, such as i-PrOH, at an elevated temperature, such as 120°C, for about 12 hours.

[0115] composition In one aspect of the present invention, a compound of Formula (I) or Formula (II) is used in combination with a therapeutic agent (e.g., a polynucleotide, a small molecule, a protein, a peptide) to be delivered to a cell or subject to form a microparticle, nanoparticle, liposome, or micelle. The agent delivered by the particle, liposome, or micelle may be in gas, liquid, or solid form, and the agent may be a polynucleotide (DNA, e.g., dsDNA, or RNA, e.g., mRNA or siRNA), a protein, a peptide, or a small molecule. The compound of Formula (I) or (II) can be combined with other lipid compounds, polymers (synthetic or natural), surfactants, cholesterol, carbohydrates, proteins, lipids, etc. to form particles. These particles can then be combined, optionally with a pharmaceutically acceptable carrier, to form a pharmaceutical composition.

[0116] Examples of pharmaceutically acceptable carriers include non-toxic (in the amounts used) solid, semi-solid, or liquid fillers, diluents, encapsulating materials, or formulations. Guidance on the formulation of small molecules, vaccines, proteins, and antibodies can be found, for example, in Remington (2020) The Science and Practice of Pharmacy 23rd Edition, D'Amico et al. (2021) Drug Deliv. and Transl. Res. 11, 353-372, and Strickley and Lambert (2021) Journal of Pharmaceutical Sciences 110:2590-2608.

[0117] The form of the pharmaceutical composition, the route of administration, the dosage and the regimen will vary depending on the condition to be treated, such as the severity of the disease, the age, weight and sex of the patient, etc. Pharmaceutical compositions can be formulated for various modes of administration, for example, topical, oral, intranasal, parenteral, intraocular, intravenous, intramuscular or subcutaneous administration.

[0118] In some embodiments, the pharmaceutical composition comprises a formulation that can be injected into a subject. Examples of components of an injectable formulation include isotonic sterile saline (e.g., monosodium or disodium phosphate, sodium chloride, potassium chloride, calcium chloride or magnesium chloride, and mixtures of such salts), buffered saline, sugar (e.g., dextrose), and water for injection. Pharmaceutical compositions include dry compositions, such as lyophilized compositions, which can be made into an injectable solution by adding sterile water or saline, as needed. The dosage used for administration can be adapted according to various parameters, such as the administration method, the pathology involved, and the duration of treatment.

[0119] Other pharmaceutically acceptable forms include tablets or other solid forms for oral administration, including time-release capsules.

[0120] How to use In another general aspect, the compounds and pharmaceutical compositions described herein are provided for treating various diseases, particularly for use in delivering therapeutic agents or therapeutic drugs (e.g., polynucleotides, small molecules, proteins, peptides) to a subject, or in cancer immunotherapy. For example, the compounds of formula (I) or (II) can be used in nanoparticle formulations to deliver DNA, RNA, or other polynucleotides to a subject or cell. For example, lipid nanoparticles (LNPs) containing compounds of formula (I) or (II) can be used to deliver RNA interference (RNAi) therapeutic agents.

[0121] As used herein, "cancer immunotherapy," also known as immuno-oncology, is a form of cancer treatment that uses the power of the body's own immune system to prevent, control, and eliminate cancer. Cancer immunotherapy comes in various forms, including targeted antibodies, cancer vaccines, adoptive cell transfer, tumor-infecting viruses, checkpoint inhibitors, cytokines, and adjuvants. mRNA is widely used in cancer immunotherapy. For example, one use of mRNA is therapeutic vaccination, which can take advantage of the ability of mRNA to deliver immune stimulants such as cytokines and chemokines.

[0122] In some embodiments, compounds of formula (II) contain ester groups and can undergo hydrolysis, allowing them to be used as biodegradable lipids. The biodegradable functionality allows these lipids to exhibit rapid clearance from plasma and tissues, thereby improving the biocompatibility of the compound and facilitating the elimination of the compound once it has served the purpose of delivering a therapeutic agent or drug to the appropriate intracellular compartment in vivo.

[0123] In certain embodiments, the compounds of formula (I) or (II) can be used in nanoparticle formulations to facilitate intracellular delivery of DNA vectors to a subject. The DNA vector can deliver a variety of different transgenes that can be expressed to provide proteins with desired activities. Examples of transgenes include those that provide healthy copies of genes in subjects with genetic defects, or new modified genes that may be useful in treating diseases or disorders, or new genes that encode proteins that have beneficial effects.

[0124] In different embodiments, the transgene encodes GAA (acid alpha-glucosidase) for the treatment of Pompe disease, TPP1 (tripeptidyl peptidase-1) for the treatment of late infantile neuronal ceroid lipofuscinosis type 2 (CLN2), ATP7B (copper transporting ATPase 2) for the treatment of Wilson's disease, alpha galactosidase for the treatment of Fabry disease, ASS1 (argininosuccinate synthase) for the treatment of citrullinemia type 1, beta-glucocerebrosidase for the treatment of Gaucher disease type 1, beta-hexosaminidase A for the treatment of Tay-Sachs disease, SERPING1 (C1 protease inhibitor or C1 esterase inhibitor) for the treatment of hereditary angioedema (HAE) (also known as C1 inhibitor deficiency types 1 and 2), or glucose-6-phosphatase for the treatment of glycogen storage disease type 1 (GSDI).

[0125] In different embodiments, the transgene is selected from the group consisting of insulin, glucagon, growth hormone (GH), parathyroid hormone (PTH), growth hormone-releasing factor (GRF), follicle-stimulating hormone (FSH), luteinizing hormone (LH), human chorionic gonadotropin (hCG), vascular endothelial growth factor (VEGF), angiopoietin, angiostatin, granulocyte colony-stimulating factor (GCSF), erythropoietin (EPO), connective tissue growth factor (CTGF), basic fibroblast growth factor (bFGF), acidic fibroblast growth factor (aFGF), epidermal growth factor (EGF), transforming growth factor (TF), and the like. The gene encoding the gene encoding transforming growth factor alpha (TGFα), platelet-derived growth factor (PDGF), insulin growth factor I or II (IGF-I or IGF-II), TGFβ, activin, bone morphogenetic protein (BMP), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin NT-3 or NT4 / 5, ciliary neurotrophic factor (CNTF), glial cell line-derived neurotrophic factor (GDNF), neurturin, agrin, netrin-1 or netrin-2, hepatocyte growth factor (HGF), ephrin, noggin, sonic hedgehog, or tyrosine hydroxylase.

[0126] In different embodiments, the transgene encodes thrombopoietin (TPO), interleukins (IL-1 through IL-36), monocyte chemotactic proteins, leukemia inhibitory factor, granulocyte-macrophage colony-stimulating factor, Fas ligand, tumor necrosis factor α or β, interferon α, β, or γ, stem cell factor, flk-2 / flt3 ligand, IgG, IgM, IgA, IgD, or IgE, chimeric immunoglobulins, antibodies, humanized antibodies, single-chain antibodies, T cell receptors, chimeric T cell receptors, single-chain T cell receptors, or class I or class II MHC molecules. For example, antibodies and immunoglobulins can be provided that target cancer cells or cells causing other diseases or disorders.

[0127] In different embodiments, the transgene is selected from the group consisting of CFTR (cystic fibrosis transmembrane conductance regulator), blood clotting (clotting) factors (Factor XIII, Factor IX (FIX), Factor VIII (FVIII), Factor X, Factor VII, Factor VIIa, or Protein C), gain-of-function blood clotting factors, erythropoietin, LDL receptor, lipoprotein lipase, ornithine transcarbamylase, β-globin, α-globin, spectrin, α-antitrypsin, adenosine deaminase, and the like. enzymes involved in lysosomal storage diseases (ARSA), hypoxanthine guanine phosphoribosyltransferase, β-25 glucocerebrosidase, sphingomyelinase, lysosomal hexosaminidase, branched-chain ketoacid dehydrogenase, hormones, growth factors, insulin-like growth factor 1 or 2, platelet-derived growth factor, epidermal growth factor, nerve growth factor, neurotrophic factor-3 and -4, brain-derived neurotrophic factor Nutritional factors, glial-derived growth factors, transforming growth factors α and β, cytokines, α-interferon, β-interferon, interferon-γ, interleukin-2, interleukin-4, interleukin-12, granulocyte-macrophage colony-stimulating factor, lymphotoxin, suicide gene products, herpes simplex virus thymidine kinase, cytosine deaminase, diphtheria toxin, cytochrome P450, deoxycytidine kinase, tumor necrosis factor, drug resistance protein proteins, tumor suppressor proteins (e.g., p53, Rb, Wt-1, NF1, von Hippel-Lindau disease (VHL), adenomatous polyposis coli (APC), peptides with immunomodulatory properties, tolerogenic or immunogenic peptides or proteins Tregitope or hCDR1, insulin, glucokinase, guanylate cyclase 2D (LCA-GUCY2D), retinal pigment epithelium-specific 65 kDa protein (RPE65), Rab escort protein 1 (total choroidal atrophy), LCA5 (LCA-lebercilin), ornithine ketoacid aminotransferase (gyrate atrophy), retinoschisin 1 (X-linked retinoschisis), X-linked retinitis pigmentosa GTPase (XLRP), MER proto-oncogene tyrosine kinase (MERTK) (autosomal recessive (AR) form of retinitis pigmentosa (RP)), ABCA4 (Stargardt), ACHM2, 3, and 4 (color blindness), anti-vascular endothelial growth factor (VEGF) polypeptides (e.g., bevacizumab, brolucizumab) , ranibizumab, aflibercept), DFNB1 (connexin 26 deafness), USH1C (Usher syndrome 1C), PKD-1 or PKD-2 (polycystic kidney disease), TPP1 (tripeptidyl peptidase-1), sulfatase, N-acetylglucosamine-1-phosphate transferase, cathepsin A, GM2-AP, NPC1, VPC2, sphingolipid activating protein, or one or more donor sequences used as a repair template for genome editing.

[0128] In different embodiments, the transgene is selected from the group consisting of erythropoietin (EPO) for the treatment of anemia, interferon alpha, interferon beta, and interferon gamma for the treatment of various immune disorders, viral infections, and cancer, interleukins (IL) and corresponding receptors, including any one of IL-1 through IL-36, for the treatment of various inflammatory diseases or immune deficiencies, chemokines, including chemokine (C-X-C motif) ligand 5 (CXCL5), for the treatment of immune disorders, granulocyte colony-stimulating factor (G-CSF) for the treatment of immune disorders such as Crohn's disease, granulocyte-macrophage colony-stimulating factor (GM-CSF) for the treatment of various human inflammatory diseases, macrophage colony-stimulating factor (M-CSF) for the treatment of various human inflammatory diseases, keratinocyte growth factor (KGF) for the treatment of epithelial tissue damage, chemokines such as monocyte chemoattractant protein-1 (MCP-1) for the treatment of recurrent miscarriage, HIV-related complications, and insulin resistance, tumor necrosis factor (TNF) and receptors for the treatment of various immune disorders, alpha 1-antitrypsin for the treatment of emphysema or chronic obstructive pulmonary disease (COPD), mucopolysaccharidosis type 1 (MPSalpha-L-iduronidase for the treatment of alpha-L-iduronidase I, ornithine transcarbamoylase (OTC) for the treatment of OTC deficiency, phenylalanine hydroxylase (PAH) or phenylalanine ammonia-lyase (PAL) for the treatment of phenylketonuria (PKU), lipoprotein lipase for the treatment of lipoprotein lipase deficiency, apolipoproteins for the treatment of apolipoprotein (Apo) AI deficiency, low-density lipoprotein receptor (LDL-R) for the treatment of familial hypercholesterolemia (FH), albumin for the treatment of hypoalbuminemia, lecithin cholesterol acyltransferase (LCAT), carbamoyl synthetase I, argininosuccinate synthetase, and Encoding argininosuccinate lyase, arginase, fumarylacetoacetate hydrolase, porphobilinogen deaminase, cystathionine beta synthase for the treatment of homocystinuria, branched-chain ketoacid decarboxylase, isovaleryl-CoA dehydrogenase, propionyl-CoA carboxylase, methylmalonyl-CoA mutase, glutaryl-CoA dehydrogenase, insulin, pyruvate carboxylase, hepatic phosphorylase, phosphorylase kinase, glycine decarboxylase, H-protein, T-protein, cystic fibrosis transmembrane conductance regulator (CFTR), ATP-binding cassette, subfamily A (ABC1), member 4 (ABCA4), or dystrophin for the treatment of Stargardt disease.

[0129] In further embodiments, the transgene encodes a protein for treating a disease or disorder selected from the group consisting of hereditary angioedema, Pompe disease, hemophilia A, hemophilia B, Fabry disease, exudative macular degeneration, Leber's hereditary optic neuropathy, and Stargardt disease. [Example]

[0130] The following examples of the present application further illustrate the nature of the present application. It should be understood that the following examples do not limit the present application, the scope of which is determined by the appended claims.

[0131] Synthesis method Unless otherwise specified, abbreviations for chemical reagents and synthetic conditions have their usual meanings known in the art, as follows: "LDA" refers to lithium diisopropylamide. "EA" refers to ethyl acetate. "PE" refers to petroleum ether. "rt" and "rt" refer to room temperature. "THF" refers to tetrahydrofuran. "DEAD" refers to diethyl azodicarboxylate. "TBAB" refers to tetrabutylammonium bromide. "DCM" refers to dichloromethane; "HOBT" refers to hydroxybenzotriazole. "LAH" refers to lithium aluminum hydride. "TLC" refers to thin layer chromatography. "Prep-TLC" refers to preparative thin layer chromatography. "TMS-I" refers to trimethylsilyl iodide. "Hex" refers to hexane. "DMF" refers to dimethylformamide. "h" or "hr" refers to hours. "min" refers to minutes. "EDCI" refers to 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide. "DMAP" refers to 4-dimethylaminopyridine. "Prep-HPLC" refers to preparative high performance liquid chromatography. "DHP" refers to dihydropyran. "DPPF" refers to 1,1'-bis(diphenylphosphino)ferrocene. "DIEA" refers to diisopropylethylamine. "TEA" refers to triethylamine. "m-CPBA" refers to meta-chloroperoxybenzoic acid. "MeOH" refers to methanol. "EtOH" refers to ethanol. "i-PrOH" refers to isopropanol. "Tol" refers to toluene.

[0132] Synthesis of compounds of formula (IA) Example 1. Synthesis of Compound SIL1-17 [ka]

[0133] Step 1. Preparation of Compounds 1-2a [ka] To a solution of compound 1-1 (2.00 g, 12.6 mmol, 1.00 equiv.) in DCM (14.0 mL) was added TEA (1.92 g, 18.9 mmol, 2.64 mL, 1.50 equiv.). Then, TsCl (2.89 g, 15.1 mmol, 1.20 equiv.) was added dropwise at 0 °C. The reaction was stirred at 20 °C for 2 h. LC-MS showed that compound 1-1 was completely consumed and the desired mass was detected. The reaction mixture was filtered, the filter cake was washed with THF (20.0 mL), and the filtrate was concentrated under reduced pressure. The crude product was used in the next step without further purification. Compound 1-2a (5.00 g, crude) was obtained as a yellow oil.

[0134] Step 2. Preparation of Compounds 1-6 [ka] To a solution of compound 1-5 (2.5 g, 13.2 mmol, 1.00 equiv.) in ACN (25.0 mL) was added K2CO3 (3.67 g, 26.5 mmol, 2.00 equiv.) and compound 1-2a (4.98 g, 15.9 mmol, 1.20 equiv.). The reaction was stirred at 90 °C for 24 h. LC-MS showed that compound 1-5 was completely consumed and the desired mass was detected. The reaction mixture was filtered, the filter cake was washed with ACN (40.0 mL), and the filtrate was concentrated under reduced pressure. The crude product was used in the next step without further purification. Compound 1-6 (3.3 g, crude) was obtained as a white solid.

[0135] Step 3. Preparation of Compounds 1-7 [ka] To a solution of compound 1-6 (3.30 g, 10.1 mmol, 1.00 equiv) in MeOH (3.00 mL) was added HCl / MeOH (15.0 mL). The reaction was stirred at 20 °C for 2 h. TLC (dichloromethane:methanol = 3:1) showed that compound 1-6 was completely consumed. The reaction mixture was concentrated under reduced pressure to remove the solvent, and the pH was adjusted to neutral with macroporous ion-base exchange resin. The crude product was used in the next step without further purification. Compound 1-7 (2.5 g, crude, 4HCl) was obtained as a white solid. 1 H NMR: 400 MHz MeOD δ 2.88 - 2.85 (m, 2 H), 2.50-2.45 (m, 4 H),2.43 - 2.40 (m, 6 H), 2.38 - 2.36 (m, 3 H), 1.29 - 2.26 (m, 3 H), 1.76 - 1.70(m, 4 H).

[0136] Step 4. Preparation of Compounds 1-8 [ka] To a solution of compound 1-7 (0.600 g, 1.31 mmol, 50% purity, 1.00 equiv.) in ACN (6.00 mL) were added K2CO3 (544 mg, 3.94 mmol, 3.00 equiv.), NaI (98.45 mg, 656 μμmol, 0.500 equiv.), and compound 1-4 (1.04 g, 2.89 mmol, 2.20 equiv.). The reaction was stirred at 90 °C for 12 h. LC-MS showed that compound 1-7 was completely consumed and the desired mass was detected. The reaction mixture was filtered, the filter cake was washed with ACN (10.0 mL), and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography (dichloromethane:methanol = 10 / 1 to 3 / 1). Compound 1-8 (0.300 g, crude) was obtained as a white solid.

[0137] Step 5. Preparation of Compounds 1-9 [ka] To a solution of compound 1-8 (0.300 g, 365 μμmol, 1.00 equiv.) in DCM (0.300 mL) was added TFA (924 mg, 8.10 mmol, 0.600 mL, 22.19 equiv.). The reaction was stirred at 20 °C for 4 h. TLC (dichloromethane:methanol = 1:1) showed that compound 1-8 was completely consumed. The reaction mixture was concentrated under reduced pressure to remove the solvent, and the pH was adjusted to neutral with macroporous ion-base exchange resin. The crude product was used in the next step without further purification. Compound 1-9 (0.130 g, crude) was obtained as a yellow oil. 1 H NMR: 400 MHz MeOD δ 3.54 - 3.51 (m, 5 H), 3.47 - 3.45 (m, 4H), 2.85 - 2.82 (m, 5 H), 2.75 - 2.72 (m, 5 H), 2.34 - 2.33 (m, 4 H), 2.33 -2.32 (m, 5 H), 2.32 - 2.29 (m, 7 H), 2.29 - 2.27 (m, 4 H), 2.20 (s, 3 H), 1.68- 1.60 (m, 5 H), 1.29 - 1.26 (m, 3 H).

[0138] Step 6. Preparation of compound SIL1-17 [ka] To a solution of compound 1-9 (0.0900 g, 223 μmol, 1.00 equiv.) in EtOH (0.5 mL), 1,2-epoxydodecane (659 mg, 3.58 mmol, 16.0 equiv.) was added. The reaction was stirred at 120 °C for 2 h. LC-MS showed that compound 1-9 was completely consumed and the desired mass was detected. The reaction was concentrated in vacuo to give the desired product. The residue was purified by silica gel chromatography (dichloromethane:methanol = 80 / 1 to 1 / 8). Compound SIL1-17 (50 mg, 39.0 μmol, 17.4% yield, 89.1% purity) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ 3.54 - 3.44 (m, 14 H), 2.82 - 2.32 (m, 32H), 1.45 - 1.72 (m, 5 H), 1.43 - 1.26 (m, 69 H), 0.90 - 0.842 (m, 12 H). LCMS:(M / 2+1):670.7

[0139] Example 2. Synthesis of compounds SIL1-7 [ka]

[0140] Step 1. Preparation of compound 2_2 [ka] To a solution of compound 2_1 (2.00 g, 9.12 mmol, 1.0 equiv.) in DCM (12.0 mL) were added DMAP (1.67 g, 13.7 mmol, 1.5 equiv.) and TsCl (2.09 g, 11.0 mmol, 1.2 equiv.). The mixture was stirred at 20 °C for 12 h. TLC (petroleum ether:ethyl acetate = 2:1) showed that compound 2_1 (2.00 g, 9.12 mmol, 1.0 equiv.) was consumed. The reaction mixture was diluted with 30 mL of brine and extracted with ethyl acetate (50 mL). The combined organic layers were dried over Na2SO4. Filtration and concentration under reduced pressure gave the residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 2 / 1). Compound 2_2 (1.60 g, 4.28 mmol, 47.0% yield) was obtained as a pale yellow oil and was checked by HNMR. 1 H NMR: (400 MHz, CDCl) δ ppm 7.78 - 7.84 (m, 2 H), 7.36 (d, J =8.00 Hz, 2 H), 4.84 (s, 1 H), 4.14 (t, J = 6.13 Hz, 2 H), 3.46 (t, J = 5.94Hz, 2 H), 3.37 - 3.41 (m, 2 H), 3.17 - 3.28 (m, 2 H), 2.46 (s, 3 H), 1.90 (m, J= 6.03 Hz, 2 H), 1.46 (s, 9 H)

[0141] Step 2. Preparation of Compound 2_7 [ka] To a solution of compound 2_6 (55.0 g, 295 mmol, 1.0 equiv.) in MeCN (330 mL) was added 1,2-dibromoethane (27.7 g, 148 mmol, 11.1 mL, 0.50 equiv.) and NaHCO3 (62.0 g, 738 mmol, 28.7 mL, 2.5 equiv.). The mixture was stirred at 90 °C for 12 h. TLC (dichloromethane:methanol = 10:1, R f =0.42), indicating that compound 2_6 was completely consumed and three new spots were formed. The reaction mixture was cooled to room temperature until complete reaction, the solvent was removed, and the residue was dissolved in ethyl acetate (500 ml). The organic solution was washed with water (300 ml) and brine (200 ml), dried over Na2SO4, and concentrated under reduced pressure. The crude product was purified by recrystallization from MeCN (150 ml) at 25 °C. Compound 2_7 (51.0 g, 128 mmol, 43.3% yield) was obtained as a white solid. 1 H NMR: (400 MHz, MeOD) δ ppm 3.43 (s, 8 H), 2.56 (s, 4 H), 2.45 -2.48 (m, 8 H), 1.46 (s, 18 H).

[0142] Step 3. Preparation of Compound 4 [ka] To a solution of compound 2_7 (10.0 g, 25.1 mmol, 1.0 equiv.) in DCM (30 mL) was added TFA (34.3 g, 301 mmol, 22.3 mL, 12 equiv.). The mixture was stirred at 20 °C for 12 h. H NMR (ET42897-1-P1A2) showed that compound 2_7 (10.0 g, 25.1 mmol, 1.0 equiv.) was consumed. The reaction mixture was concentrated under reduced pressure to give a residue. Compound 2_8 was dissociated with alkaline resin to a pH of about 7-8 for filtration and rotary evaporation. Compound 2_8 (6.00 g, crude) was obtained as a pale yellow solid. 1 It was checked by HNMR. 1H NMR: (400 MHz, DO) δ ppm 3.36 - 3.46 (m, 8 H), 3.14 - 3.30 (m,12 H).

[0143] Step 4. Preparation of Compound 2_9 [ka] To a solution of compound 2_8 (2.43 g, 12.2 mmol, 2.2 equiv.) in ACN (12 mL) was added K2CO3 (1.15 g, 8.35 mmol, 1.5 equiv.) and 2-[2-(tert-butoxycarbonylamino)ethoxy]ethyl 4-methylbenzenesulfonate (2 g, 5.56 mmol, 1.0 equiv.). The mixture was stirred at 90 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed that compound 2_8 (2.43 g, 12.2 mmol, 2.2 equiv.) and 2-[2-(tert-butoxycarbonylamino)ethoxy]ethyl 4-methylbenzenesulfonate (2.00 g, 5.56 mmol, 1.0 equiv.) were consumed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 20 / 1 to 5 / 1) to give compound 2_9 (1.2 g) as a white oil. It was checked by HNMR. 1 H NMR: (400 MHz, MeOD) δ ppm 3.61 (s, 2 H), 3.47 (s, 2 H), 3.34(d, J = 3.63 Hz, 2 H), 3.18 - 3.25 (m, 1 H), 3.21 (s, 1 H), 3.12 (s, 4 H), 2.45- 2.83 (m, 15 H), 1.44 (s, 9 H).

[0144] Step 5. Preparation of Compound 2_3 [ka] To a solution of compound 2_2 (959 mg, 2.57 mmol, 1.1 equiv.) in ACN (6.0 mL) were added NaI (175 mg, 1.17 mmol, 0.50 equiv.), Cs2CO3 (1.14 g, 3.50 mmol, 1.5 equiv.), and compound 2_9 (900 mg, 2.33 mmol, 1.0 equiv.). The mixture was stirred at 90 °C for 6 h. TLC (petroleum ether:ethyl acetate = 1:1) showed that compound 2_9 (900 mg, 2.33 mmol, 1.0 equiv.) had been consumed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. This was combined with the purified products ET42897-18, 21, and 24. The residue was purified by preparative HPLC (column: Phenomenex Gemini-NX C18 75 × 30 mm × 3 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 5% to 35%, 12 min). Compound 2_3 (TFA) (500 mg, 23.0% yield) was obtained as a white oil and checked by LCMS.

[0145] Step 6. Preparation of Compound 2_4 [ka] To a solution of compound 2_3 (500 mg, 713 μmol, 1.0 equiv., TFA) in DCM (5.0 mL) was added TFA (1.22 g, 10.7 mmol, 792 μL, 15 equiv.). The mixture was stirred at 20 °C for 3 h. HNMR (ET42897-27-P1A2) showed that compound 2_3 (500 mg, 713 μmol, 1.0 equiv., TFA) was consumed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The crude product was dissociated with alkaline resin to a pH of about 7-8 for filtration and rotary evaporation. Compound 2_4 (0.28 g, crude) was obtained as a yellow oil. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, MeOD) δ ppm 3.61 (t, J = 5.57 Hz, 2 H), 3.43 -3.53 (m, 6 H), 2.77 (td, J = 5.28, 3.19 Hz, 4 H), 2.42 - 2.65 (m, 20 H), 1.73 -1.84 (m, 2 H).

[0146] Step 7. Preparation of compounds SIL1-7 [ka] To a solution of compound 2_4 (0.25 g, 671.06 μmol, 1.0 equiv.) in EtOH (5.0 mL), 2-decyloxirane (1.24 g, 6.71 mmol, 10 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42897-29-P1A1) showed that compound 2_4 (0.25 g, 671.06 μmol, 1.0 equiv.) was consumed, and the product RT = 0.704 min. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was combined with the purified product of ET42897-28 and purified by column chromatography (SiO2, dichloromethane:methanol = 20 / 1 to 10 / 1). Compound SIL1-7 (60 mg, 7% yield) was obtained as a yellow oil. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, CDCl) δ ppm 3.39 - 3.68 (m, 11 H), 2.27 - 2.91(m, 34 H), 1.82 (s, 2 H), 1.15 - 1.55 (m, 77 H), 0.89 (t, J = 6.48 Hz, 13 H)

[0147] Example 3. Synthesis of compounds SIL1-8 [ka]

[0148] Step 1. Preparation of compound 5_2 [ka] To a solution of compound 10_1 (25 g, 141 mmol, 1.0 equiv.) and compound A (23.52 g, 169 mmol, 15.27 mL, 1.2 equiv.) in EtOH (150 mL) was added NaOH (2 M, 141.03 mL, 2.0 equiv.). The mixture was stirred at 25° C. for 3 hours. TLC (petroleum ether:ethyl acetate=1:1, product R f =0.43), indicating the reaction of the raw materials was complete. HO (100 mL) was added to the mixture. The aqueous phase was extracted with MTBE (500 mL, 200 mL). The combined organic phase was washed with brine (100 mL), dried over anhydrous NaSO, filtered, and concentrated in vacuo. The residue was purified by column chromatography (SiO, petroleum ether:ethyl acetate = 100:1 to 2:1). Compound 5_2 (24 g, 101 mmol, 72.3% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 4.89 (br s, 1H), 3.76 (t, J = 5.9 Hz,2H), 3.28 - 3.39 (m, 2H), 2.67 (q, J = 6.7 Hz, 4H), 1.82 - 1.90 (m, 2H), 1.46(s, 9H).

[0149] Step 2. Preparation of compound 5_3 [ka] To a solution of compound 5_2 (15.4 g, 65.4 mmol, 1.0 equiv.), DMAP (15.9 g, 130 mmol, 2.0 equiv.), and DIEA (16.9 g, 130 mmol, 22.8 mL, 2.0 equiv.) in DCM (92 mL) was added TsCl (13.7 g, 71.9 mmol, 1.1 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 h. TLC (petroleum ether:ethyl acetate=2:1, product R f=0.59), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 50:1 to 5:1). Compound 5_3 (8.54 g, 33.6 mmol, 51.4% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 4.91 (br s, 1H), 3.66 (t, J = 6.3 Hz,2H), 3.33 (br d, J = 5.9 Hz, 2H), 2.68 (td, J = 6.8, 15.7 Hz, 4H), 1.98 - 2.09(m, 2H), 1.45 (s, 9H)

[0150] Step 3. Preparation of compound 5_4 [ka] To a solution of compound 2_8 (11.72 g, 59.10 mmol, 2.5 equiv.) and compound 5_3 (6 g, 23.64 mmol, 1 equiv.) in ACN (30 mL) was added NaI (1.77 g, 11.8 mmol, 0.5 equiv.) and K2CO3 (4.90 g, 35.46 mmol, 1.5 equiv.). The mixture was stirred at 90 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed the product R f =0.38), indicating the reaction of the raw materials was complete. The mixture was filtered, and the filtrate was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 40:1). Compound 5_4 (5 g, 12.03 mmol, 50.88% yield) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ = 4.95 (br dd, J = 3.0, 6.1 Hz, 1H), 3.26- 3.35 (m, 2H), 2.89 (t, J = 4.9 Hz, 4H), 2.37 - 2.67 (m, 22H), 1.76 (m, J =7.3 Hz, 2H), 1.44 (s, 9H)

[0151] Step 4. Preparation of Compound 5_5 [ka] To a solution of compound 5_4 (4.1 g, 9.86 mmol, 1 equiv.) and compound 10_3 (3.55 g, 14.80 mmol, 1.5 equiv.) in ACN (24 mL) was added K2CO3 (2.05 g, 14.80 mmol, 1.5 equiv.) and NaI (739 mg, 4.93 mmol, 0.5 equiv.). The mixture was stirred at 90 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed the product R f =0.43), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, ethyl acetate:methanol = 100:1 to 20:1). Compound 5_5 (4.2 g, crude) was obtained as a yellow oil.

[0152] Step 5. Preparation of Compound 5_6 [ka] To a solution of compound 5_5 (800 mg, 1.29 mmol, 1 equiv.) in DCM (10 mL) was added TFA (7.70 g, 67.5 mmol, 5 mL, 52.3 equiv.). The mixture was stirred at 20° C. for 4 h. LCMS showed that the starting material was completely consumed. The mixture was evaporated to dryness. Compound 5_6 (450 mg, crude) was obtained as a yellow oil.

[0153] Step 6. Preparation of compound SIL8 [ka] Two batches. To a solution of compound 5_6 (225 mg, 537 μmol, 1 equiv.) in EtOH (15 mL), 2-decyloxirane (990 mg, 5.37 mmol, 10 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42415-63-P1A1, product: RT = 0.703 min). The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, ethyl acetate:methanol = 100:1 to 40:1). Compound SIL1-8 (70 mg, 60.55 μmol, 5.63% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 3.58 - 3.67 (m, 4H), 2.26 - 2.85 (m,45H), 1.72 - 1.82 (m, 2H), 1.21 - 1.51 (m, 77H), 0.86 - 0.91 (m, 12H).

[0154] Example 4. Synthesis of compounds SIL1-9 [ka]

[0155] Step 1. Preparation of compound 10_2 [ka] To a solution of compound 10_1 (25 g, 141.03 mmol, 1 equiv.) and compound B (21.2 g, 169 mmol, 12.0 mL, 1.2 equiv.) in EtOH (150 mL) was added NaOH (2 M, 141.03 mL, 2 equiv.). The mixture was stirred at 25° C. for 3 hours. TLC (petroleum ether:ethyl acetate=1:1) showed that the product R f=0.43), indicating the reaction of the raw materials was complete. HO (100 mL) was added to the mixture. The aqueous phase was extracted with MTBE (500 mL, 200 mL). The combined organic phase was washed with brine (100 mL), dried over anhydrous NaSO, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (SiO, petroleum ether:ethyl acetate = 100:1 to 2:1). Compound 10_2 (19 g, 85.8 mmol, 60.8% yield) was obtained as a yellow oil. 1 H NMR: 400MHz CDCl3 δ = 4.80 - 4.99 (m, 1H), 3.76 (t, J = 5.9Hz, 2H), 3.34 (br t, J = 6.4 Hz, 2H), 2.76 (t, J = 5.9 Hz, 2H), 2.67 (t, J =6.6 Hz, 2H), 2.23 (br d, J = 4.1 Hz, 1H), 1.46 (s, 9H).

[0156] Step 2. Preparation of compound 10_3 [ka] To a solution of compound 10_2 (5 g, 22.5 mmol, 1 equiv.), DMAP (5.52 g, 45.1 mmol, 2 equiv.), and TEA (4.57 g, 45.2 mmol, 6.29 mL, 2 equiv.) in DCM (30 mL) was added TosCl (4.74 g, 24.8 mmol, 1.1 equiv.) at 0 °C. The mixture was stirred at 0-25 °C for 3 h. TLC (petroleum ether:ethyl acetate = 2:1) revealed the presence of product R. f =0.53), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 50:1 to 5:1). Compound 10_3 (2.5 g, 10.4 mmol, 46.1% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 4.91 (br s, 1H), 3.61 - 3.68 (m, 2H),3.28 - 3.37 (m, 2H), 2.85 - 2.92 (m, 2H), 2.70 (t, J = 6.5 Hz, 2H), 1.45 (s,9H).

[0157] Step 3. Preparation of compound 10_6 [ka] To a solution of compound 10_5 (400 mg, 4.64 mmol, 1 equiv.) and compound 10_3 (2.34 g, 9.75 mmol, 2.1 equiv.) in ACN (20 mL) was added NaI (348 mg, 2.32 mmol, 0.5 equiv.) and K2CO3 (2.57 g, 18.5 mmol, 4 equiv.). The mixture was stirred at 90 °C for 2 h. TLC showed that approximately 10% of compound 10_3 remained, and one new major spot with greater polarity was detected. The mixture was filtered, and the filtrate was evaporated in vacuo. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 40:1). Compound 10_6 (900 mg, 1.83 mmol, 39.3% yield) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ = 5.03 (br s, 2H), 3.32 (q, J = 5.8 Hz,4H), 2.47 - 2.73 (m, 20H), 1.45 (s, 18H)

[0158] Step 4. Preparation of compound 10_7 [ka] To a solution of compound 10_6 (900 mg, 1.83 mmol, 1 equiv.) in MeOH (27 mL), HCl / MeOH (4 M, 27.00 mL, 59.1 equiv.) was added. The mixture was stirred at 25° C. for 2 hours. TLC (dichloromethane:methanol=10:1, product R f=0.04) indicated the reaction of the raw materials was complete. The mixture was filtered with MTBE (20 mL) and the filter cake was evaporated in vacuo to give compound 10_7 (600 mg, crude) as a white solid. 1 H NMR: 400 MHz DMSO-d6 δ = 2.97 (t, J = 7.2 Hz, 2H), 2.59 - 2.66(m, 2H), 2.53 - 2.58 (m, 4H), 2.31 - 2.48 (m, 13H).

[0159] Step 5. Preparation of compounds SIL1-9 [ka] A solution of compound 10_7 (300 mg, 1.03 mmol, 1 equiv.) and 2-decyloxirane (1.13 g, 6.15 mmol, 6 equiv.) in EtOH (20 mL) was prepared. The mixture was stirred at 120 °C for 12 h. LCMS (ET42415-26-P1A1, product: RT = 0.737 min). The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 40:1). Compound SIL1-9 (70 mg, 60.5 μmol, yield 5.90%, purity 89%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 3.62 (br d, J = 3.6 Hz, 4H), 3.17 -3.28 (m, 3H), 2.31 - 2.85 (m, 30H), 1.25 - 1.51 (m, 73H), 0.89 (t, J = 6.7 Hz,12H)

[0160] Example 5. Synthesis of Compounds SIL1-10 [ka]

[0161] Step 1. Preparation of compound 11-2 [ka] To a solution of compound 11-1 (0.10 g, 1.16 mmol, 1.00 equiv.) in DMF (0.60 mL) were added K2CO3 (401 mg, 2.90 mmol, 2.50 equiv.) and compound 1-4 (918 mg, 2.55 mmol, 2.20 equiv.). The reaction was stirred at 110 °C for 12 h. LC-MS showed that compound 11-1 was completely consumed and the desired mass was detected. The residue was poured into water (2.00 mL). The aqueous phase was extracted with DCM (5.00 mL). The combined organic phase was dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The residue was purified by silica gel chromatography (dichloromethane:methanol = 30 / 1). Compound 11-2 (0.26 g, 564.47 μmol, 48.62% yield) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ 5.17 (s, 2 H), 3.60 - 3.56 (m, 4H), 3.54- 3.49 (m, 4 H), 3.31 - 3.29 (m, 4 H), 2.59 - 2.41 (m, 11 H), 1.45 (s, 18 H)

[0162] Step 2. Preparation of compound 11-3 [ka] To a solution of compound 11-2 (0.260 g, 564 μmol, 1.00 equiv) in MeOH (0.300 mL) was added HCl / MeOH (4 M, 1.50 mL). The reaction was stirred at 10 °C for 4 h. TLC (dichloromethane:methanol = 10:1) showed that compound 11-2 was completely consumed. The reaction was concentrated under reduced pressure, and the pH was adjusted to alkaline with 6 M K2CO3 (aq). The crude product was used in the next step without further purification. The pH of the solution was adjusted to alkaline. Compound 11-3 (0.158 g, crude) was obtained as a white solid. 1H NMR: 400 MHz DMSO δ 2.78 - 2.66 (m, 10 H),2.63 - 2.65 (m, 5H), 2.14 - 2.05 (m, 4 H), 2.04 - 2.00 (m, 4 H).

[0163] Step 3. Preparation of Compounds SIL1-10 [ka] To a solution of compound 11-3 (158 mg, 606 μmol, 1.00 equiv.) in EtOH (4.00 mL), 2-decyloxirane (671 mg, 3.64 mmol, 6.00 equiv.) was added. The reaction was stirred at 120° C. for 60 h. The mixture was stirred at 15° C. for 16 h. TLC (dichloromethane:methanol = 10 / 1) showed that compound 11-3 was completely consumed. The residue was purified by silica gel chromatography (dichloromethane:methanol = 80 / 1, 20 / 1). Compound SIL1-10 (50.0 mg, 47.9 μmol, 7.90% yield, 95.7% purity) was obtained as a gray oil. 1 H NMR: 400 MHz CDCl δ 3.50 - 3.80 (m, 16 H), 2.75 - 2.84 (m, 2H), 2.53 - 2.64 (m, 10 H), 2.18 - 2.51 (m, 7 H), 1.26 - 1.44 (m, 105 H), 0.89(t, J = 6.4 Hz, 12H). LCMS:(M / 2+1):499.6

[0164] Example 6. Synthesis of Compounds SIL1-11 [ka]

[0165] Step 1. Preparation of compound 12_1 [ka] To a solution of compound 13_1 (1 g, 6.98 mmol, 1 eq.) and compound 10_3 (3.52 g, 14.6 mmol, 2.1 eq.) in DMF (50 mL) was added NaI (523 mg, 3.49 mmol, 0.5 eq.) and K2CO3 (3.86 g, 27.9 mmol, 4 eq.). The mixture was stirred at 100 °C for 12 h. TLC (dichloromethane:methanol = 8:1, product R f =0.28), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 20:1). Compound 12_1 (400 mg, 688 μmol, yield 9.86%, purity 94.6%) was obtained as a yellow oil.

[0166] Step 2. Preparation of compound 12_2 [ka] To a solution of compound 12_1 (400 mg, 727 μmol, 1 equiv.) in MeOH (4 mL), HCl / MeOH (4 M, 4 mL, 21.9 equiv.) was added. The mixture was stirred at 25° C. for 2 hours. TLC (dichloromethane:methanol=10:1, product R f =0.02) indicated that the reaction of the raw materials was complete. The mixture was evaporated to dryness. Compound 12_2 (230 mg, crude) was obtained as a yellow oil.

[0167] Step 3. Preparation of Compound SIL1-11 [ka] To a solution of compound 12_2 (100 mg, 286 μmol, 1 equiv.) in EtOH (2 mL), 2-decyloxirane (527 mg, 2.86 mmol, 10 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42415-42-P1A1, product: RT = 0.713 min). The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, ethyl acetate:methanol = 100:1 to 40:1). Compound SIL1-11 (70 mg, 62.9 μmol, yield 22.00%, purity 97.7%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 3.58 - 3.74 (m, 4H), 2.28 - 2.90 (m,37H), 1.51 - 1.52 (m, 1H), 1.25 - 1.51 (m, 73H), 0.92 (t, J = 6.8 Hz, 12H).

[0168] Example 7. Synthesis of compounds SIL1-12 [ka]

[0169] Step 1. Preparation of compound 12_1 [ka] To a solution of compound 13_1 (1 g, 6.98 mmol, 1 eq.) and compound 10_3 (3.52 g, 14.6 mmol, 2.1 eq.) in DMF (50 mL) was added NaI (523 mg, 3.49 mmol, 0.5 eq.) and K2CO3 (3.86 g, 27.9 mmol, 4 eq.). The mixture was stirred at 100 °C for 12 h. TLC (dichloromethane:methanol = 8:1, product R f=0.28), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 20:1). Compound 12_1 (400 mg, 688 μmol, yield 9.86%, purity 94.6%) was obtained as a yellow oil.

[0170] Step 2. Preparation of compound 12_2 [ka] To a solution of compound 12_1 (400 mg, 727 μmol, 1 equiv.) in MeOH (4 mL), HCl / MeOH (4 M, 4 mL, 21.9 equiv.) was added. The mixture was stirred at 25° C. for 2 hours. TLC (dichloromethane:methanol=10:1, product R f =0.02) indicated that the reaction of the raw materials was complete. The mixture was evaporated to dryness. Compound 12_2 (230 mg, crude) was obtained as a yellow oil.

[0171] Step 3. Preparation of Compound SIL1-12 [ka] To a solution of compound 13-3 (560 mg, 1.76 mmol, 1 equiv.) in EtOH (11 mL), 2-decyloxirane (1.95 g, 10.6 mmol, 6 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42513-15-P1A1, product RT = 0.690 min) indicated the reaction was complete. The reaction was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100 / 1 to 0 / 1) to give compound SIL1-12 (200 mg, 190 μmol, 10.8% yield) as a yellow oil. LCMS: [M+H] + :m / z=1054.0 1 H NMR: (400 MHz CD3OD) δ=2.62-2.82 (m, 7H), 2.54-2.58 (m, 16H),2.30 - 2.50 (m, 3H), 2.16 (s, 1H), 1.30 (m, 75H), 0.89-0.92 (m, 12H)

[0172] Example 8. Synthesis of compounds SIL1-13 [ka]

[0173] Step 1. Preparation of compound 17_1 [ka] To a solution of compound 18_1 (1 g, 6.36 mmol, 1 eq.) and compound 17_1 (3.20 g, 13.3 mmol, 2.1 eq.) in DMF (50 mL) was added NaI (476 mg, 3.18 mmol, 0.5 eq.) and K2CO3 (3.52 g, 25.4 mmol, 4 eq.). The mixture was stirred at 100 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed the product R f =0.36), indicating the reaction of the raw materials was complete. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 40:1). Compound 17_1 (400 mg, 695 μmol, yield 10.9%, purity 98%) was obtained as a yellow oil.

[0174] Step 2. Preparation of compound 17_2 [ka] To a solution of compound 17_1 (400 mg, 709 μmol, 1 equiv.) in MeOH (10 mL) was added HCl / MeOH (4 M, 10 mL, 56.3 equiv.). The mixture was stirred at 25° C. for 2 h. LCMS showed that the starting material was completely consumed. The mixture was evaporated to dryness. Compound 17_2 (290 mg, crude) was obtained as a yellow oil.

[0175] Step 3. Preparation of Compound SIL1-13 [ka] To a solution of compound 17_2 (130 mg, 357 μmol, 1 equiv.) in EtOH (13 mL), 2-decyloxirane (658 mg, 3.58 mmol, 10 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42415-43-P1A1, product: RT = 0.712 min). The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, ethyl acetate:methanol = 100:1 to 40:1). Compound SIL1-13 (70 mg, 60.3 μmol, yield 8.44%, purity 94.9%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ = 3.57 - 3.69 (m, 4H), 2.28 - 2.86 (m,35H), 1.17 - 1.53 (m, 78H), 0.89 (t, J = 6.7 Hz, 12H) LCMS:ET42415-43-P1A1

[0176] Example 9. Synthesis of Compound SIL1-16 [ka]

[0177] Step 1. Preparation of compound 18-2 [ka] A solution of compound 18-1 (450 mg, 2.86 mmol, 1 equiv.), compound 1-4 (2.16 g, 6.01 mmol, 2.1 equiv.), and K2CO3 (1.58 g, 11.5 mmol, 4 equiv.) in DMF (4.5 mL) was degassed and purged with N2 three times. The mixture was then stirred at 90 °C under a N2 atmosphere for 12 h. LCMS (ET42513-10-P1A2, product RT = 0.051 min) indicated the reaction was complete. The reaction was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100 / 1 to 0 / 1) to give compound 18-2 (760 mg, 1.43 mmol, 50.0% yield) as a yellow oil. LCMS: [M+H] + :m / z=532.0 1 H NMR: (400 MHz CDCl) δ=3.49-3.59 (m, 13H), 3.29-3.35 (m, 6H),2.36 (s, 3H), 1.44 (m, 22H)

[0178] Step 2. Preparation of compound 18-3 [ka] To a solution of compound 18-2 (760 mg, 1.43 mmol, 1 equiv.) in MeOH (11 mL), HCl / MeOH (4 M, 357 μL, 1 equiv.) was added. The mixture was stirred at 25 °C for 4 h. LCMS (ET42513-12-P1A1, product RT = 0.056 min) showed the reaction was complete. The pH value of the reaction solution was adjusted to 7-8 with saturated aqueous K2CO3 solution. The crude product was used directly in the next step without further purification. Compound 18-3 (810 mg, crude) was obtained as a yellow oil. LCMS: [M+H] + :m / z=332.0

[0179] Step 3. Preparation of compound SIL1-16 [ka] To a solution of compound 18-3 (810 mg, 2.44 mmol, 1 equiv.) in EtOH (8 mL), 2-decyloxirane (2.70 g, 14.7 mmol, 6 equiv.) was added. The mixture was stirred at 120 °C for 12 h. LCMS (ET42513-13-P1A1, product RT = 0.699 min) indicated the reaction was complete. The reaction was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100 / 1 to 0 / 1). The crude product was purified by reverse-phase HPLC (column: Phenomenex Luna C18 100 × 30 mm × 5 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 35% to 70%, 10 min) to give compound SIL1-16 (40 mg, 37.4 μmol, yield 1.53%) as a yellow oil. LCMS: [M+H] + :m / z=1068.0 1 H NMR: (400 MHz CD3OD) δ= 2.31 (s, 3H), 2.16(s, 1H), 1.30 - 1.48(m, 76H), 0.89-0.92 (m, 12H) Other compounds such as SIL1-1, SIL1-2, SIL1-3, SIL1-4, SIL1-5, SIL1-6, SIL1-14, SIL1-15, and SIL1-16 were also prepared in a similar manner as above.

[0180] Synthesis of compounds of formula (IB) Example 1. Synthesis of compound SIL2-1 [ka]

[0181] Step 1. Preparation of Compounds 1-2 [ka] Compound 1a (3.44 g, 24.64 mmol, 1.1 equiv., HCl), EDCI (5.15 g, 26.88 mmol, 1.2 equiv.), HOBt (3.63 g, 26.88 mmol, 1.2 equiv.), and TEA (4.53 g, 44.80 mmol, 6.24 mL, 2 equiv.) were added to a solution of compound 1-1 (5 g, 22.40 mmol, 1 equiv.) in DCM (30 mL) at 0° C. The mixture was stirred at 20° C. for 12 hours. TLC (petroleum ether:ethyl acetate=2:1, R f =0.1) indicated that compound 1-1 was completely consumed and many new spots were formed. The reaction mixture was washed with water (30 mL) and NaHCO3 (50 mL). The organic phase was separated, washed with brine (30 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The crude product was used in the next step without further purification. Compound 1-2 (7.5 g, crude) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ 7.27 - 7.40 (m, 5 H) 6.70 (br s, 1 H)5.39 (br s, 1 H) 5.06 - 5.17 (m, 2 H) 4.49 - 4.61 (m, 1 H) 4.22 - 4.35 (m, 1 H)3.72 (s, 3 H) 1.38 (d, J = 7.00 Hz, 6 H).

[0182] Step 2. Preparation of Compounds 1-3a [ka] To a solution of compound 1-2 (7 g, 22.70 mmol, 1 equiv.) in MeOH (70 mL) was added Pd / C (2 g, 1.69 mmol, 10% purity) under N. The suspension was degassed under vacuum and purged with H several times. The mixture was stirred under H (30 psi) at 50 °C for 12 h. TLC (petroleum ether:ethyl acetate = 2:1, R f= 0.00) indicated that compound 1-2 was completely consumed and many new spots were formed. After filtration through Celite, the filter cake was washed with MeOH, and the filtrate was concentrated. The crude product was used in the next step without further purification. Compound 1-3a (4.2 g, crude) was obtained as a gray solid.

[0183] Step 3. Preparation of Compounds 1-3 [ka] Compound 1-3a (4.2 g, 24.11 mmol, 1 equiv.) was dissolved in toluene (39 mL) and then stirred at 130° C. for 12 hours. 1 HNMR showed that the starting material was completely consumed. The suspension mixture was cooled to 0 °C. The solid was filtered and washed with petroleum ether (20 mL × 2). The crude product was used in the next step without further purification. Compound 1-3 (2.7 g, 18.99 mmol, 78.78% yield) was obtained as a gray solid. 1 H NMR: 400 MHz DMSO-d6 δ 8.09 (br s, 2 H) 3.78 - 3.94 (m, 2 H)1.26 (d, J = 7.00 Hz, 6 H).

[0184] Step 4. Preparation of compound C1 [ka] Compound 1-3 (2.2 g, 15.48 mmol, 1 equiv.) was added portionwise to a solution of LiAlH4 (1.76 g, 46.43 mmol, 3 equiv.) in THF (22 mL), and the reaction mixture was stirred at 70° C. for 12 h. 1HNMR showed that the starting material was completely consumed. The reaction mixture was quenched by adding 1.8 mL of water at 0 ° C., then diluted with 1.8 mL of 15% NaOH solution and 5.4 mL of water, filtered, the filter cake was washed with THF, and the filtrate was concentrated under reduced pressure to give a residue. Compound C1 (2 g, crude) was obtained as a yellow solid. 1 H NMR: 400 MHz MeOD δ 2.90 (dd, J = 12.26, 2.75 Hz, 2 H) 2.63 -2.74 (m, 2 H) 2.37 (dd, J = 12.19, 10.69 Hz, 2 H) 1.03 (d, J = 6.38 Hz, 6 H).

[0185] Step 5. Preparation of Compounds 1-4 [ka] To a solution of compound 3a (1.89 g, 7.88 mmol, 1.5 equiv.) in MeCN (30 mL), K2CO3 (2.18 g, 15.76 mmol, 3 equiv.), NaI (393.80 mg, 2.63 mmol, 0.5 equiv.), and compound C1 (600 mg, 5.25 mmol, 1 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS (ET54476-78-P1A1, product: RT = 0.503 min) showed that the starting material was completely consumed. TLC (dichloromethane:methanol = 10:1, R f =0.30) indicated that compound C1 was completely consumed and many new spots were formed. The reaction mixture was filtered, and the filter cake was washed with ACN. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 20 / 1 to 0 / 1). Compound 1-4 (1.2 g, 2.30 mmol, 43.85% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.97 (br s, 2 H) 3.32 (br d, J = 5.88 Hz,4 H) 3.02 - 3.08 (m, 2 H) 2.78 (br d, J = 8.88 Hz, 2 H) 2.64 - 2.69 (m, 8 H)2.39 - 2.55 (m, 4 H) 2.06 - 2.17 (m, 2 H) 1.45 (s, 18 H) 1.07 (br d, J = 4.25Hz, 6 H)

[0186] Step 6. Preparation of Compounds 1-5 [ka] To a solution of compound 1-4 (1.2 g, 2.30 mmol, 1 equiv.) in DCM (1 mL) was added TFA (3.08 g, 27.01 mmol, 2.00 mL, 11.72 equiv.). The mixture was stirred at 20° C. for 12 hours. TLC (dichloromethane:methanol=10:1, R f =0.00) indicated that compound 1-4 was completely consumed and one new spot was formed. The reaction mixture was concentrated. The crude product was used in the next step without further purification. Compound 1-5 (1.9 g, crude, TFA) was obtained as a brown oil.

[0187] Step 7. Preparation of compound SIL2-1 [ka] Compound 1-5 (0.5 g, 1.15 mmol, 1 equiv., TFA) was dissolved in methanol, adjusted to pH 8-9 with alkaline resin, filtered, and concentrated to give 200 mg of residue. To a solution of the residue in EtOH (10 mL) was added 2-decyloxirane (1.70 g, 9.20 mmol, 8 equiv.). The mixture was stirred at 120 °C for 72 h. TLC (dichloromethane:methanol = 10:1, R f=0.42) indicated that compound 1-5 was completely consumed and many new spots were formed. The reaction mixture was concentrated. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100 / 1 to 10 / 1). Compound SIL2-1 (70 mg, 66.17 μmol, yield 5.75%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 3.58 - 3.74 (m, 4 H) 2.97 - 3.17 (m, 4 H)2.16 - 2.96 (m, 30 H) 1.25 - 1.48 (m, 72 H) 1.14 (br s, 6 H) 0.89 (t, J = 6.82Hz, 12 H)

[0188] Example 2. Synthesis of compound SIL2-2 [ka]

[0189] Step 1. Preparation of Compound 2-2 [ka] To a solution of compound 2-1 (0.9 g, 6.33 mmol, 1.0 equiv.) in THF (90 mL) was added LAH (721 mg, 19.0 mmol, 3.0 equiv.) at 20° C. The mixture was stirred at 78° C. for 12 hours. TLC (dichloromethane:methanol=10:1) showed that the reactants were consumed and the product R f = 0.0. The reaction mixture was quenched by adding 0.72 mL of H2O, 0.72 mL of NaOH (15%), 2.16 mL of H2O at 0 °C, added Na2SO4, filtered, and concentrated under reduced pressure to give a residue. No purification was performed. Compound 2-2 (0.7 g, crude) was obtained as a white solid and checked by HNMR. 1 H NMR: (400 MHz MeOD) δ ppm 2.75 - 2.93 (m, 4 H), 2.61 - 2.70 (m,2 H), 1.15 (d, J = 6.75 Hz, 6 H).

[0190] Step 2. Preparation of Compounds 2-3 [ka] To a solution of compound 3a (4.17 g, 13.9 mmol, 3.0 equiv.) in ACN (10 mL), K2CO3 (1.92 g, 13.9 mmol, 3.0 equiv.), NaI (348 mg, 2.32 mmol, 0.5 equiv.), and compound 2-2 (0.53 g, 4.64 mmol, 1.0 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS showed that the reactant was consumed and the product RT = 0.658 min. The reaction mixture was filtered, and the filter cake was washed with ACN. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 100 / 1 to 3 / 1). Compound 2-3 (0.3 g, 576 μmol, 12.4% yield) was obtained as a pale yellow solid and characterized by HNMR. 1 H NMR:ET54385-43-P1 (400 MHz, CDCl3) δ ppm 4.89 - 5.06 (m, 2 H), 3.25 - 3.39 (m,4 H), 1.46 (s, 18 H), 2.39 - 2.74 (m, 16 H), 1.04 (d, J = 6.13 Hz, 5 H).

[0191] Step 3. Preparation of Compounds 2-4 [ka] To a solution of compound 2-3 (0.3 g, 576 μmol, 1.0 equiv.) in DCM (10 mL) was added TFA (2.31 g, 20.3 mmol, 1.5 mL, 35 equiv.). The mixture was stirred at 20° C. for 12 h. LCMS showed the reactant was consumed and the product RT=0.044 min. The reaction mixture was concentrated under reduced pressure to remove DCM (10 mL). The crude product was dissolved in MeOH (50 mL) and adjusted to pH=7 by adding alkaline resin (NH4HCO3). No purification was performed. Compound 2-4 was obtained as a brown oil and checked by HNMR. 1 H NMR: 400 MHz, CDCl δ ppm 2.89 - 2.97 (m, 4 H), 2.44 - 2.81 (m,19 H), 2.01 - 2.28 (m, 9 H), 1.06 (d, J = 6.38 Hz, 6 H).

[0192] Step 4. Preparation of compound SIL2-2 [ka] To a solution of compound 2-4 (0.16 g, 499 μmol, 1.0 equiv.) in EtOH (16 mL) was added compound 2A (736 mg, 3.99 mmol, 8.0 equiv.). The mixture was stirred at 120 °C for 60 h. LCMS showed the reactant was consumed and the product RT = 0.349 min. The reaction mixture was concentrated under reduced pressure to remove EtOH (16 mL). The residue was purified by column chromatography (SiO, petroleum ether / ethyl acetate = 5 / 1 to DCM:MeOH = 30 / 1).

[0193] Compound SIL2-2 (52.5 mg, 47.3 μmol, 9.47% yield) was obtained as a pale yellow oil and checked by HNMR. 1 H NMR: (400 MHz, CDCl) δ ppm 3.76 (s, 1 H), 3.63 (s, 4 H), 3.63 (s, 4 H), 3.21 (s, 3 H),2.30 - 2.89 (m, 25 H), 1.86 (t, J = 6.54, 3.30 Hz, 1 H), 1.19 - 1.54 (m, 68 H),1.04 (s, 6 H), 0.89 (t, J = 6.75 Hz, 12 H).

[0194] Example 3. Synthesis of Compound SIL2-3 [ka]

[0195] Step 1. Preparation of Compound 3-2 [ka] To a solution of compound 3-1 (8.50 g, 41.8 mmol, 1.0 equiv.) and compound 1a (10.6 g, 92.0 mmol, 2.2 equiv.) in DCM (60.0 mL) was added EDCI (14.4 g, 75.3 mmol, 1.8 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed complete consumption of the reactants and the formation of many new spots. The residue was poured into saturated sodium bicarbonate (10.0 mL) and stirred for 5 min. The aqueous phase was extracted with DCM (15.0 mL × 2). The combined organic phase was washed with brine (10.0 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. No purification was performed. Compound 3-2 (17.5 g, crude) was obtained as a yellow oil.

[0196] Step 2. Preparation of Compound 3-3 [ka] To a solution of compound 3-2 (17.5 g, 58.4 mmol, 1.0 equiv) in DCM (30.0 mL) was added TFA (21.3 mL, 288 mmol, 4.93 equiv) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC showed that reactant 1 was completely consumed and many new spots had formed. It was filtered and concentrated in vacuo. No purification was performed. Compound 3-3 (26.2 g, crude) was obtained as a yellow oil.

[0197] Step 3. Preparation of Compounds 3-4 [ka] To a solution of compound 3-3 (26.2 g, 131 mmol, 1.0 equiv) in DCM (160 mL) was added pyridine (64.1 g, 810 mmol, 65.4 mL, 6.20 equiv) at 0 °C. The mixture was stirred at 20 °C for 12 h. HNMR indicated that the reaction was complete and the product was the desired product. It was filtered and concentrated in vacuo. The crude product was stirred with ethyl acetate at 20 °C for 1 h. Compound 3-4 (1.50 g, crude) was obtained as a white solid and checked by HNMR. 1 H NMR: (400 MHz, DMSO-d6) δ 8.08 (s, 2H), 3.81 (m, 2H), 1.68 (m, 4H),0.84 (m, 6H).

[0198] Step 4. Preparation of Compounds 3-5 [ka] To a solution of compound 3-4 (1.50 g, 8.81 mmol, 1.0 equiv.) in THF (15.0 mL) was added LAH (1.00 g, 26.4 mmol, 3.0 equiv.). The mixture was stirred at 75 °C for 12 h. LCMS showed that reactant 1 was completely consumed and one major peak with the desired mass was detected. The mixture was cooled to 5 °C. Next, water (1.00 mL), NaOH-water (1.00 mL), and then water (3.00 mL) were added to the mixture and stirred for 5 min. The mixture was filtered and concentrated in vacuo. No purification was performed. Compound 3-5 (1.00 g, crude) was obtained as a yellow oil.

[0199] Step 5. Preparation of Compounds 3-6 [ka] To a solution of tert-butyl N-[2-(2-chloroethylsulfanyl)ethyl]carbamate (1.75 g, 7.31 mmol, 1.3 equiv.) in ACN (15.0 mL), KCO (2.33 g, 16.9 mmol, 3 equiv.), NaI (422 mg, 2.81 mmol, 0.5 equiv.), and compound 3-5 (800 mg, 5.62 mmol, 1.0 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS showed the reactants were consumed and the product RT = 0.550 min. The reaction mixture was filtered, and the filter cake was washed with ACN. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 100 / 1, 0 / 1). Compound 3-6 (900 mg, 1.64 mmol, 29.2% yield) was obtained as a yellow oil.

[0200] Step 6. Preparation of Compounds 3-7 [ka] To a solution of compound 3-6 (300 mg, 547 μmol, 1.0 equiv) in DCM (10.0 mL) was added TFA (693 mg, 6.08 mmol, 450 μL, 11.1 equiv) at 0° C. The mixture was stirred at 20° C. for 12 hours. LCMS showed that the reactants were consumed. The mixture was filtered and concentrated in vacuo. Compound 3-7 (180 mg, crude) was obtained as a yellow oil without further purification.

[0201] Step 7. Preparation of compounds SIL2-3 [ka] To a solution of compound 3-7 (180 mg, 516 μmol, 1.0 equiv.) in EtOH (20.0 mL), 2-decyloxirane (761 mg, 4.13 mmol, 8.0 equiv.) was added. The mixture was stirred at 120 °C for 72 h. LC-MS showed that compound 3-7 was completely consumed. It was filtered and concentrated in vacuo. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, DCM / MeOH = 100 / 1, 0 / 1). Compound SIL2-3 (160 mg, 147 μmol, 28.5% yield) was obtained as a yellow oil. 1 It was checked by HNMR. 1 H NMR: (400 MHz, chloroform-d) δ 3.65 (s, 4H), 3.25 (s, 3H), 2.75 (m, 6H),2.60 (m, 12H), 2.35 (m, 9H), 1.45 (m, 80H), 0.89 (m,18H).

[0202] Compounds SIL2-4 to SIL2-8 were prepared in the same manner as above. Their structures and 1H-NMR data are summarized in Table 1. In addition, compound SIL2-9 can also be prepared in the same manner as above. [Table 1] TIFF2025531749000114.tif226149 TIFF2025531749000115.tif226149

[0203] Example 4. Synthesis of compound SIL2-10 [ka]

[0204] Step 1. Preparation of compound 10-2 [ka] To a solution of compound 10-1 (10.0 g, 37.7 mmol, 1.0 equiv.) in THF (60 mL), DCC (9.33 g, 45.2 mmol, 9.15 mL, 1.2 equiv.) and compound 1a (5.21 g, 45.2 mmol, 1.2 equiv.) were added at 0° C. The mixture was stirred at 20° C. for 12 hours. TLC (dichloromethane:methanol=10:1) showed that the product R f =0.60) indicated that the reactants were exhausted. The reaction mixture was filtered, the filter cake was washed with THF, and the combined organic layers were concentrated under reduced pressure to give a residue. No purification was carried out. Compound 2 (14.0 g, crude) was obtained as a white solid.

[0205] Step 2. Preparation of compound 10-3 [ka] To a solution of compound 10-2 (14.0 g, 38.6 mmol, 1.0 equiv.) in DCM (20 mL) was added TFA (32.3 g, 284 mmol, 21.0 mL, 7.34 equiv.). The mixture was stirred at 20° C. for 12 hours. TLC (dichloromethane:methanol=10:1, product R f=0.43) indicated that the reactants were exhausted. The reaction mixture was concentrated under reduced pressure to remove DCM (20 mL). The crude was dissolved in Tol (100 mL) to remove TFA twice. No purification was performed. Compound 10-3 (18.0 g, crude) was obtained as a yellow oil.

[0206] Step 3. Preparation of compound 10-4 [ka] To a solution of compound 10-3 (10.1 g, 38.6 mmol, 1.0 equiv) in DCM (30 mL) was added pyridine (24.5 g, 310 mmol, 25.0 mL, 8.0 equiv). The mixture was stirred at 20 °C for 12 h. LCMS showed the reactant was consumed and the product RT = 0.592 min. The reaction mixture was concentrated under reduced pressure to remove DCM (30 mL). The crude product was triturated with MeOH (50 mL) at 25 °C for 60 min. Compound 10-4 (0.82 g, 2.79 mmol, 14.4% yield) was obtained as a white solid and checked by HNMR. 1 H NMR: (400 MHz, DMSO-d6) δ ppm 7.92 (s, 2 H), 7.11 - 7.36 (m, 7 H),7.03 (d, J = 7.25 Hz, 4 H), 3.97 (s, 2 H), 2.57 (dd, J = 13.51, 4.63 Hz, 2 H),2.23 (dd, J = 13.51, 6.13Hz, 2H).

[0207] Step 4. Preparation of compound 10-5 [ka] To a solution of compound 10-4 (1.60 g, 5.44 mmol, 1.0 equiv.) in THF (80 mL) was added LAH (619 mg, 16.3 mmol, 3.0 equiv.) at 20 °C. The mixture was stirred at 78 °C for 12 h. LCMS showed that the reactants were consumed and the product RT = 0.612 min. The reaction mixture was quenched by adding 0.62 mL of HO, 0.62 mL of NaOH (15%), and 1.86 mL of HO at 0 °C, followed by addition of NaSO, filtration, and concentration under reduced pressure to give a residue. The residue was purified by column chromatography (SiO, petroleum ether / ethyl acetate = 100 / 1 to 3 / 1). Compound 10-5 (0.93 g, 3.49 mmol, 64.2% yield) was obtained as a pale yellow solid. 1 It was checked by HNMR. 1 H NMR: (400 MHz, DMSO-d6) δ ppm 7.11 - 7.35 (m, 8 H), 3.33 (s, 2 H),3.17 (s, 1 H), 2.72 - 2.86 (m, 4 H), 2.53 - 2.71 (m, 4 H).

[0208] Step 5. Preparation of compound 10-6 [ka] To a solution of compound 3a (2.09 g, 8.73 mmol, 2.5 equiv.) in ACN (30 mL), K2CO3 (1.45 g, 10.5 mmol, 3.0 equiv.) and NaI (262 mg, 1.75 mmol, 0.5 equiv.), compound 10-5 (0.93 g, 3.49 mmol, 1.0 equiv.) was added. The mixture was stirred at 90 °C for 12 h. LCMS showed that the reactant was consumed and the product RT = 1.001 min. The reaction mixture was filtered, and the filter cake was washed with ACN. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 100 / 1 to 2 / 1). Compound 10-6 (1.60 g, 2.38 mmol, 68.1% yield) was obtained as a yellow oil and characterized by HNMR. 1H NMR: (400 MHz, CDCl) δ ppm 7.16 - 7.40 (m, 9 H), 4.95 (s, 2 H),3.18 - 3.46 (m, 4 H), 2.51 - 3.06 (m, 18 H), 2.35 (d, J = 11.25 Hz, 2 H), 1.48(s, 18 H).

[0209] Step 6. Preparation of compound 10-7 [ka] To a solution of compound 10-6 (1.60 g, 2.38 mmol, 1.0 equiv) in DCM (10 mL) was added TFA (7.70 g, 67.5 mmol, 5 mL, 28.4 equiv). The mixture was stirred at 20 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed that the reactants were consumed. The reaction mixture was concentrated under reduced pressure to remove DCM (10 mL). The crude product was dissolved in MeOH (50 mL) and adjusted to pH = 7 by adding alkaline resin (NH4HCO3). No purification was performed. Compound 10-7 (0.5 g, 1.06 mmol, 44.5% yield) was obtained as a brown oil and checked by HNMR. 1 H NMR: (400 MHz, CDCl) δ ppm 7.29 (br d, J=7.00 Hz, 3 H), 7.17 -7.25 (m, 5 H), 5.78 - 6.24 (m, 2 H), 2.46 - 3.02 (m, 24 H).

[0210] Step 7. Preparation of compound SIL2-10 [ka] To a solution of compound 10-7 (0.5 g, 1.06 mmol, 1.0 equiv) in EtOH (50 mL) was added 2-decyloxirane (1.56 g, 8.46 mmol, 8.0 equiv). The mixture was stirred at 120 °C for 72 h. LCMS showed the reactant was consumed and the product RT = 0.918 min. The reaction mixture was concentrated under reduced pressure to remove EtOH (50 mL). The residue was purified by column chromatography (SiO, petroleum ether / ethyl acetate = 5 / 1 to DCM:MeOH = 30 / 1).

[0211] Compound SIL2-10 (52.25 mg, 43.2 μmol, yield 4.08%) was obtained as a yellow oil. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, CDCl) δ ppm 7.27 - 7.32 (m, 3 H), 7.17 - 7.24 (m,4 H), 3.54 - 3.84 (m, 5 H), 2.24 - 3.02 (m, 26 H), 1.17 - 1.57 (m, 72 H), 0.89(t, J = 6.82 Hz, 12H).

[0212] Example 5. Synthesis of compound SIL2-11 [ka]

[0213] Step 1. Preparation of compound 11-2 [ka] To a solution of compound 11-1 (5.00 g, 19.4 mmol, 1.0 equiv.) and compound 1a (4.92 g, 42.8 mmol, 2.2 equiv.) in DCM (30.0 mL) was added EDCI (6.70 g, 35.0 mmol, 1.8 equiv.) at 0 °C. The mixture was stirred at 25 °C for 16 h. TLC (dichloromethane:methanol = 10:1) showed complete consumption of the reactants and the formation of many new spots. The residue was poured into saturated sodium bicarbonate (5.00 mL) and stirred for 5 min. The aqueous phase was extracted with DCM (10.0 mL × 2). The combined organic phase was washed with brine (5.00 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. No purification was performed. Compound 11-2 (7.95 g, crude) was obtained as a white solid.

[0214] Step 2. Preparation of compound 11-3 [ka] To a solution of compound 11-2 (7.95 g, 22.4 mmol, 1.0 equiv) in DCM (20.0 mL) was added TFA (9.54 mL, 129 mmol, 5.74 equiv) at 0 °C. The mixture was stirred at 25 °C for 2 h. TLC showed that reactant 1 was completely consumed and many new spots had formed. It was filtered and concentrated in vacuo. No purification was performed. Compound 11-3 (12.7 g, crude) was obtained as a yellow oil.

[0215] Step 3. Preparation of compound 11-4 [ka] To a solution of compound 11-3 (12.7 g, 50.1 mmol, 1.0 equiv) in DCM (80.0 mL) was added pyridine (31.2 g, 394 mmol, 31.8 mL, 7.88 equiv) at 0 °C. The mixture was stirred at 20 °C for 12 h. HNMR showed that the reaction was complete and the product was the desired product. It was filtered and concentrated in vacuo. The crude product was stirred with ethyl acetate at 20 °C for 1 h. Compound 11-4 (1.70 g, crude) was obtained as a white solid.1 Checked by 1 H NMR. 1 H NMR: (400 MHz, DMSO- d6 ) δ 7.95 (s, 1H), 3.62 (s, 1H), 3.25 (s, 2H),2.76 (m, 2H), 1.55 (m, 10H), 1.05 (m, 10H).

[0216] Step 4. Preparation of compound 11-5 [ka] To a solution of compound 11-4 (2.00 g, 7.18 mmol, 1.0 equiv) in THF (20.0 mL) was added LAH (872 mg, 23.0 mmol, 3.2 equiv). The mixture was stirred at 75 °C for 12 h. LCMS showed that reactant 1 was completely consumed and one major peak with the desired mass was detected. The mixture was cooled to 5 °C. Next, water (0.90 mL), NaOH-water (0.90 mL), and then water (2.70 mL) were added to the mixture and stirred for 5 min. The mixture was filtered and concentrated in vacuo. No purification was performed. Compound 11-5 (1.40 g, crude) was obtained as a white oil.

[0217] Step 5. Preparation of compound 11-6 [ka] To a solution of tert-butyl N-[2-(2-chloroethylsulfanyl)ethyl]carbamate (2.14 g, 8.94 mmol, 1.6 equiv.) in ACN (20.0 mL), KCO (2.32 g, 16.8 mmol, 3 equiv.) and NaI (419 mg, 2.80 mmol, 0.5 equiv.), compound 11-5 (1.40 g, 5.59 mmol, 1.0 equiv.) was added. The mixture was stirred at 90 °C for 12 h. LCMS showed the reactants were consumed and the product RT = 0.649 min. The reaction mixture was filtered, and the filter cake was washed with ACN.

[0218] The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 100 / 1, 0 / 1) to give compound 11-6 (400 mg, 609 μmol, yield 10.9%) as a yellow oil.

[0219] Step 6. Preparation of compound 11-7 [ka] To a solution of compound 11-6 (370 mg, 563 μmol, 1.0 equiv) in DCM (6.00 mL) was added TFA (855 mg, 7.50 mmol, 555 μL, 13.3 equiv). The mixture was stirred at 20 °C for 12 h. LCMS showed the reactants were exhausted. It was filtered and concentrated in vacuo. No purification was performed. Compound 11-7 (250 mg, crude) was obtained as a yellow oil.

[0220] Step 7. Preparation of compound SIL2-11 [ka] To a solution of compound 11-7 (150 mg, 328 μmol, 1.0 equiv.) in EtOH (40.0 mL), 2-decyloxirane (605 mg, 3.28 mmol, 10 equiv.) was added. The mixture was stirred at 120 °C for 72 h. LC-MS showed that compound 11-7 was completely consumed. It was filtered and concentrated in vacuo. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, DCM / MeOH = 100 / 1, 0 / 1). Compound SIL2-11 (120 mg, 101 μmol, 30.6% yield) was obtained as a yellow oil. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, CDCl) δ 3.65 (m, 4H), 3.45 (m, 1H), 3.25 (m, 2H),2.55 (m, 16H), 2.45 (m, 5H), 2.10 (s, 1H), 1.95 (s, 1H), 1.55 (m, 10H), 1.35(m, 71H), 0.89 (m,12H).

[0221] Example 6. Synthesis of compound SIL2-12 [ka]

[0222] Step 1. Preparation of Compound D [ka] Compound 12-1 (3.00 g, 11.1 mmol, 1.00 equiv) was added to a solution of HCl / MeOH (4.00 M, 21.0 mL, 1.00 equiv) at 0 °C. The mixture was stirred at 80 °C for 8 h. LCMS (ET54505-120-P1A1, RT = 0.357 min) showed the desired product. Concentration under reduced pressure gave the residue, which was not further purified. Compound D (1.84 g, 9.93 mmol, 89.8% yield) was obtained as an off-white solid. 1 H NMR: (400 MHz, CDCl) δ ppm 8.88 (s, 2 H), 4.10 (s, 1 H), 3.83(s, 3 H), 3.50 (s, 1 H), 1.55 - 2.12 (m, 10 H), 0.85 - 1.44 (m, 5 H).

[0223] Step 2. Preparation of compound 12-A [ka] To a solution of compound 12-1 (2.50 g, 9.21 mmol, 1.00 equiv.) in DCM (20.0 mL) at 0 °C, HOBt (1.49 g, 11.1 mmol, 1.20 equiv.) and EDCI (2.12 g, 11.1 mmol, 1.20 equiv.) were added. After 30 min, compound D (1.88 g, 10.1 mmol, 1.10 equiv.) and DIEA (1.49 g, 11.5 mmol, 2.01 mL, 1.25 equiv.) were added. The reaction was stirred at 25 °C for 7 h. TLC (dichloromethane:methanol = 10:1) showed that the material had been consumed. The DCM solution was washed with 1 N HCl, saturated NaHCO3, and brine, dried over MgSO4, evaporated in vacuo, and was not further purified. Compound 12-A (3.30 g, 7.52 mmol, 81.7% yield) was obtained as a brown oil. 1 H NMR: (400 MHz, CDCl) δ ppm 6.25-6.50 (m, 1 H), 4.78 - 4.92 (m, 1H), 4.64 (m, 1 H), 4.13 (m, 1 H), 3.73 (s, 3 H), 1.67 (m, 11 H), 1.49 (s, 9 H),1.19 (m, 7) H), 0.80 - 1.04 (m, 4 H).

[0224] Step 3. Preparation of compound 12-B [ka] Compound 12-A (3.30 g, 7.52 mmol, 1.00 equiv.) was dissolved in DCM (5.00 mL). TFA (6.16 g, 54.0 mmol, 4.00 mL, 7.18 equiv.) was added to the mixture at 20° C. The mixture was stirred for 12 hours. TLC (petroleum ether:ethyl acetate=0:1, material R f =0.42, product R f =0.00) indicated that the material had been consumed. Concentration under reduced pressure gave a residue. No further purification was carried out. Compound 12-B (3.96 g, crude) was obtained as a brown oil.

[0225] Step 3. Preparation of compound 12-C [ka] Compound 12-B (3.96 g, 11.7 mmol, 1.00 equiv) was dissolved in Tol (30.0 mL), degassed, and the mixture was purged with N2 three times. TEA (5.92 g, 58.5 mmol, 8.14 mL, 5.00 equiv) was added to the mixture. The mixture was stirred at 120 °C for 12 h. TLC (dichloromethane:methanol = 5:1, material rf = 0.31, dichloromethane:methanol = 20:1, product rf = 0.33) showed that the material had been consumed. Concentration under reduced pressure gave a residue. The crude product was triturated with HO at 20 °C for 30 min. No further purification was performed. Compound 12-C (1.45 g, 4.73 mmol, 40.4% yield) was obtained as a white solid. 1 H NMR: (400 MHz, DMSO) δ ppm 8.17 (s, 2 H), 3.75 (s, 2 H), 1.45 -1.65 (m, 16 H), 1.15 - 1.24 (m, 6 H), 0.81 - 0.91 (m, 4 H).

[0226] Step 5. Preparation of compound 12-5 [ka] Compound 12-C (1.00 g, 3.26 mmol, 1.00 equiv) was dissolved in THF (30.0 mL) at 25 °C. LAH (619 mg, 16.3 mmol, 5.00 equiv) was added to the reaction at 25 °C. The reaction was stirred at 75 °C for 12 h. TLC (dichloromethane:methanol = 5:1, material rf = 0.63) showed material remained. 0.6 mL of HO was added to the reaction at 0 °C, and the mixture was poured with 0.6 mL of 15% aqueous NaOH and 1.80 mL of HO. Anhydrous sodium sulfate was added to the reaction. Filtration and concentration under reduced pressure gave a residue. No further purification was performed. Compound 12-5 (1.20 g, crude) was obtained as an off-white solid.

[0227] Step 6. Preparation of compound 12-6 [ka] Compound 3a (781 mg, 3.26 mmol, 1.81 equiv) was dissolved in MeCN (30.0 mL), and KCO (744 mg, 5.39 mmol, 3.00 equiv) and NaI (135 mg, 898 μmol, 0.5 equiv) were added to the mixture at 20 °C. Compound 12-5 (0.5 g, 1.80 mmol, 1.00 equiv) was added to the mixture at 20 °C. The reaction was stirred at 90 °C for 12 h. TLC (dichloromethane:methanol = 5:1, petroleum ether:ethyl acetate = 0:1, material rf = 0.03, product rf = 0.52) and LCMS (ET54505-129-P1A1, RT = 0.669 min) indicated the desired product. Concentration under reduced pressure gave the residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 0 / 1) to give compound 12-6 (0.629 g, 584 μmol, yield 32.5%) as a brown solid.

[0228] Step 7. Preparation of compound 12-7 [ka] Compound 12-6 (0.629 g, 918 μmol, 1.00 equiv) was dissolved in DCM (3.00 mL). TFA (1.45 g, 12.7 mmol, 944 μL, 14.0 equiv) was added to the mixture at 20 °C. The mixture was stirred for 12 h. TLC (petroleum ether:ethyl acetate = 0:1, material rf = 0.42, product rf = 0.00) showed that the material was consumed. Concentration under reduced pressure gave a residue. Resin (5.00 g) was added to adjust the pH to 7-9. No further purification was performed. Compound 12-7 (0.40 g, 825 μmol, 89.8% yield) was obtained as a brown oil.

[0229] Step 8. Preparation of compound SIL2-12 [ka] Compound 12-7 (0.124 g, 256 μmol, 1.00 equiv) was dissolved in EtOH (40.0 mL). 2-Decyloxirane (471 mg, 2.56 mmol, 10.0 equiv) was added to the mixture. The mixture was stirred at 120 °C for 144 h. TLC (dichloromethane:methanol = 10:1, product rf = 0.49) indicated that the material had been consumed. Concentration under reduced pressure gave a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 0 / 1). Compound SIL2-12 (0.05 g, 40.9 μmol, 16.0% yield) was obtained as a brown oil. 1 H NMR: (400 MHz, CDCl) δ ppm 3.20 - 3.85 (m, 13 H), 2.23 - 2.92(m, 23 H), 1.26- 1.76 (m, 100 H), 0.85- 0.95 (m, 12 H). LCMS: (MS / 2+H=611, RT=3.432 min)

[0230] Example 7. Synthesis of compound SIL2-13 [ka]

[0231] Step 1 Preparation of compound 13a [ka] Compound 13a-1 (1 g, 8.69 mmol, 1 equiv.) was dissolved in MeOH (10 mL) and cooled to 0 °C, followed by the addition of TMSCl (2.57 g, 23.64 mmol, 3 mL, 2.72 equiv.). The mixture was warmed to 20 °C and stirred for 12 h. LCMS (ET54476-35-P1A1, product: RT = 0.048 min) showed complete consumption of the starting material. The solvent was removed under vacuum. The resulting yellow oil was dissolved in hot EtOAc and precipitated with cyclohexane. Compound 13a (1.2 g, 7.25 mmol, 83.42% yield, HCl) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ 8.71 (br s, 3 H) 5.86 (ddt, J = 17.01,9.91, 7.24, 7.24 Hz, 1 H) 5.20 - 5.38 (m, 2 H) 4.23 - 4.33 (m, 1 H) 3.80 (s, 3H) 2.79 - 2.91 (m, 2H)

[0232] Step 2. Preparation of compound 13-2 [ka] To a solution of compound 13-1 (1 g, 4.65 mmol, 1 equiv.) in DCM (6 mL), HOBt (753.30 mg, 5.58 mmol, 1.2 equiv.), TEA (940.23 mg, 9.29 mmol, 1.29 mL, 2 equiv.), EDCI (1.07 g, 5.58 mmol, 1.2 equiv.), and compound 13a (846.38 mg, 5.11 mmol, 1.1 equiv., HCl) were added. The mixture was stirred at 15 °C for 12 h. TLC (petroleum ether:ethyl acetate = 2:1, R f=0.24) indicated that compound 13-1 was completely consumed and two new spots were formed. The reaction mixture was washed with water (10 mL), the organic phase was separated, and the aqueous phase was extracted with ethyl acetate (20 mL), washed with NaHCO (30 mL), brine (30 mL), dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. Compound 13-2 (1.3 g, crude) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.64 (br d, J = 7.25 Hz, 1 H) 5.59 - 5.83(m, 2 H) 5.07 - 5.21 (m, 4 H) 4.99 (br s, 1 H) 4.62 - 4.71 (m, 1 H) 4.09 - 4.23(m, 1 H) 3.74 (s, 3 H) 2.43 - 2.66 (m, 4 H) 1.45 (s, 9 H)

[0233] Step 3. Preparation of compound 13-3 [ka] To a solution of compound 13-2 (9.7 g, 29.72 mmol, 1 equiv.) in DCM (15 mL) was added TFA (49.30 g, 432.34 mmol, 32.01 mL, 14.55 equiv.), and the mixture was stirred at 20° C. for 12 h. TLC (petroleum ether:ethyl acetate=2:1, R f =0.00) indicated that compound 13-2 was completely consumed and one new spot was formed. The reaction mixture was concentrated under reduced pressure. The crude product was used in the next step without further purification. Compound 13-3 (14.7 g, crude, TFA) was obtained as a yellow oil.

[0234] Step 4. Preparation of compound 13-4 [ka] To a solution of compound 13-3 (14.7 g, 64.97 mmol, 1 equiv.) in toluene (147 mL) was added TEA (32.87 g, 324.83 mmol, 45.21 mL, 5 equiv.), and the mixture was stirred at 130° C. for 12 h. 1 HNMR showed that the starting material was completely consumed. The reaction mixture was diluted with ethyl acetate (10 mL) and filtered. The filter cake was concentrated. The crude product was used in the next step without further purification. Compound 13-4 (3.6 g, 18.53 mmol, 28.53% yield) was obtained as a white solid. 1 H NMR: 400 MHz DMSO-d6 δ 8.02 - 8.15 (m, 2 H) 5.71 (ddt, J =16.96, 10.27, 7.08, 7.08 Hz, 2 H) 5.06 - 5.15 (m, 4 H) 3.83 - 3.97 (m, 2 H)2.34 - 2.48 (m, 4 H)

[0235] Step 5. Preparation of compound C13 [ka] Compound 13-4 (3.6 g, 18.53 mmol, 1 equiv.) was added portionwise to a solution of LiAlH (3.52 g, 92.67 mmol, 5 equiv.) in THF (60 mL) at 20 °C, and the reaction mixture was stirred at 70 °C for 12 h. LCMS (ET54476-64-P1A1, product: RT = 0.044 min) showed that the starting material was completely consumed. The reaction mixture was cooled to 0 °C and quenched by the dropwise addition of 3.5 mL of water at 0 °C. It was then diluted with 3.5 mL of 15% NaOH solution and 10.5 mL of water, filtered, and concentrated under reduced pressure to give a residue. Compound C13 (3.6 g, crude) was obtained as a white solid. 1 H NMR: 400 MHz CDCl δ 5.70 - 5.84 (m, 2 H) 5.04 - 5.15 (m, 5 H)2.81 - 2.88 (m, 2 H) 2.70 - 2.81 (m, 4 H) 2.29 - 2.40 (m, 2 H) 2.18 - 2.26 (m,2 H) 1.68 (dt, J = 5.94, 2.78 Hz, 2H)

[0236] Step 6. Preparation of compound 13-5 [ka] To a solution of compound 3a (1.30 g, 5.41 mmol, 1.5 equiv.) in MeCN (30 mL), K2CO3 (1.50 g, 10.83 mmol, 3 equiv.), NaI (270.46 mg, 1.80 mmol, 0.5 equiv.), and compound C13 (600 mg, 3.61 mmol, 1 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS (ET54476-69-P1A1, product: RT = 0.572 min) showed that the starting material was completely consumed. TLC (dichloromethane:methanol = 10:1, R f =0.50) indicated that compound C13 was completely consumed and many new spots were formed. The reaction mixture was filtered, and the filter cake was washed with ACN. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 20 / 1 to 0 / 1). Compound 13-5 (880 mg, 1.54 mmol, 42.57% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 5.78 (ddt, J = 16.93, 9.80, 7.33, 7.33Hz, 2 H) 5.01 - 5.13 (m, 4 H) 4.95 (br d, J = 0.88 Hz, 2 H) 3.25 - 3.37 (m, 4H) 2.73 - 2.87 (m, 2 H) 2.51 - 2.70 (m, 13 H) 2.11 - 2.47 (m, 7 H) 1.45 (s, 18H)

[0237] Step 7. Preparation of compound 13-6 [ka] To a solution of compound 13-5 (1.1 g, 1.92 mmol, 1 equiv.) in DCM (0.5 mL) was added TFA (2.93 g, 25.66 mmol, 1.9 mL, 13.36 equiv.). The mixture was stirred at 20° C. for 12 h. TLC (dichloromethane:methanol=10:1, R f =0.00) indicated that compound 13-5 was completely consumed and one new spot was formed. The reaction mixture was concentrated. The crude product was used in the next step without further purification. Compound 13-6 (1.9 g, crude, TFA) was obtained as a brown oil.

[0238] Step 7. Preparation of compound SIL2-13 [ka] Compound 13-6 (500 mg, 1.03 mmol, 1 equiv., TFA) was dissolved in methanol, adjusted to pH 8-9 with alkaline resin, filtered, and concentrated to give 200 mg of residue. To a solution of the residue in EtOH (4 mL) was added compound 2a (1.51 g, 8.22 mmol, 8 equiv.). The mixture was stirred at 120 °C for 72 h. LCMS (ET54476-92-P1A1, product: RT = 0.953 min) showed that the starting material was completely consumed. TLC (dichloromethane:methanol = 10:1, R f =0.44) showed that many new spots were formed. The reaction mixture was concentrated. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100 / 1 to 10 / 1). Compound SIL2-13 (100 mg, 90.10 μmol, 8.77% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 5.69 - 5.86 (m, 2 H) 4.98 - 5.16 (m, 4 H)3.63 (br s, 4 H) 3.01 - 3.39 (m, 4 H) 2.52 - 2.98 (m, 22 H) 2.28 - 2.51 (m, 10H) 2.11 - 2.27 (m, 2 H) 1.25 - 1.48 (m, 72 H) 0.89 (t, J = 6.75 Hz, 12 H)

[0239] Example 8. Synthesis of compound SIL2-15 [ka]

[0240] Step 1. Preparation of compound 1b [ka] To a solution of compound 1a (4.50 g, 28.9 mmol, 1.0 equiv., HCl) in MeOH (30 mL) was added HCl / MeOH (4 M, 45 mL, 6.2 equiv.). The mixture was stirred at 80° C. for 12 h. LCMS showed the reactant was consumed and the product RT=0.044 min. The reaction mixture was concentrated under reduced pressure to remove MeOH (30 mL). No purification was performed. Compound 1b (6.00 g, crude, 2HCl) was obtained as a white solid. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, MeOD) δ ppm 4.25 - 4.33 (m, 1 H), 3.83 - 3.86 (m,3 H), 3.73 - 3.82 (m, 1 H), 3.38 - 3.41 (m, 3 H), 3.33 - 3.37 (m, 1 H).

[0241] Step 2. Preparation of compound 15-2 [ka] To a solution of compound 15-1 (5.50 g, 25.1 mmol, 1.0 equiv.) in DCM (24 mL), EDCI (5.77 g, 30.1 mmol, 1.2 equiv.) and HOBt (4.07 g, 30.1 mmol, 1.2 equiv.), compound 1b (4.08 g, 27.6 mmol, 90% purity, 1.1 equiv.), and TEA (7.62 g, 75.3 mmol, 10.5 mL, 3.0 equiv.) were added. The mixture was stirred at 20 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed that the reactants were consumed. The reaction mixture was quenched by adding 50 mL of HO and extracted with 100 mL of DCM. The organic layer was washed with 50 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. No purification was performed. Compound 15-2 (6.00 g, crude) was obtained as a pale yellow oil and checked by HNMR. 1 H NMR: (400 MHz, CDCl) δ ppm 5.36 - 5.47 (m, 1 H), 4.65 - 4.75 (m,1 H), 4.20 - 4.37 (m, 1 H), 3.79 - 3.88 (m, 2 H), 3.72 - 3.79 (m, 3 H), 3.57 -3.64 (m, 1 H), 3.46 - 3.54 (m, 1 H), 3.38 - 3.43 (m, 3 H), 3.31 - 3.36 (m, 3H), 1.38 - 1.52 (m, 9 H).

[0242] Step 3. Preparation of compound 15-3 [ka] To a solution of compound 15-3 (6.00 g, 17.9 mmol, 1.0 equiv.) in DCM (12 mL) was added TFA (13.9 g, 122 mmol, 9.00 mL, 6.8 equiv.). The mixture was stirred at 20° C. for 12 hours. TLC (dichloromethane:methanol=10:1, product R f= 0.0) indicated that the reactants were exhausted. The reaction mixture was concentrated under reduced pressure to remove DCM (12 mL). No purification was carried out. Compound 15-3 (14.5 g, crude, TFA) was obtained as a pale yellow oil.

[0243] Step 4. Preparation of compound 15-4 [ka] To a solution of methyl compound 15-3 (14.5 g, 61.9 mmol, 1.0 equiv.) in ToL (87 mL) was added TEA (31.3 g, 310 mmol, 43 mL, 5.0 equiv.), and the mixture was stirred at 130° C. for 12 h. 1 H NMR showed that the reactants were consumed. The reaction mixture was concentrated under reduced pressure to remove Tol (87 mL). The crude product was triturated with ethyl acetate (50 mL) at 25 °C for 60 min. Compound 15-4 (2.6 g, 12.9 mmol, 41.5% yield) was obtained as a brown solid. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, DMSO) δ ppm 8.03 - 8.15 (m, 2 H), 3.86 (d, J =1.63 Hz, 2 H), 3.64 - 3.71 (m, 2 H), 3.44 (dd, J = 9.63, 2.75 Hz, 2 H), 3.20 -3.25 (m, 6 H)

[0244] Step 5. Preparation of compound C15 [ka] To a suspension of compound 15-4 (2.60 g, 12.9 mmol, 1.0 equiv) in THF (52 mL) at 20 °C, LAH (1.46 g, 38.6 mmol, 3.0 equiv) was added to the mixture at 20 °C for 2 min. The reaction solution was then heated to 70 °C and stirred for 12 h. LCMS showed the reactant was consumed and the product RT = 0.044 min. The reaction was cooled to 0 °C, and water (1.46 mL) was added dropwise to the reaction solution under N2, generating a large amount of foam. Then, 15% aqueous NaOH (1.46 mL) was added dropwise at 0 °C. After 5 min, water (4.38 mL) was added dropwise to the reaction solution at 0 °C. Na2SO4 was then added to the mixture, which was then warmed to 25 °C and stirred for 15 min. The mixture was filtered, and the filter cake was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 100 / 1 to 1 / 1). Compound C15 (1.10 g, 6.31 mmol, 49.1% yield) was obtained as a white solid. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, DMSO-d6) δ ppm 3.20 - 3.25 (m, 6 H), 3.10 - 3.20 (m,4 H), 2.77 - 2.84 (m, 2 H), 2.60 - 2.70 (m, 2 H), 2.22 (t, J = 10.69 Hz, 2 H)

[0245] Step 6. Preparation of compound 15-5 [ka] To a solution of compound 3a (2.61 g, 10.9 mmol, 1.9 equiv.) in ACN (20 mL), K2CO3 (2.38 g, 17.2 mmol, 3.0 equiv.), NaI (430 mg, 2.87 mmol, 0.5 equiv.), and compound C15 (1.0 g, 5.74 mmol, 1.0 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS showed that the reactants were consumed and the product RT = 0.515 min. The reaction mixture was filtered, and the filter cake was washed with ACN. It was combined with the purified ET54385-77. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 100 / 1 to 2 / 1). Compound 15-5 (0.8 g, 22% yield) was obtained as a yellow solid. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, CDCl) δ ppm 4.99 (s, 2 H), 3.38 - 3.51 (m, 4 H), 3.23- 3.37 (m, 10 H), 2.80 - 3.05 (m, 4 H), 2.51 - 2.76 (m, 12 H), 2.31 t, J =10.15 Hz, 2 H), 1.39 - 1.54 (m, 18 H).

[0246] Step 7. General Procedure for the Preparation of Compound 15-6 [ka] To a solution of compound 15-5 (0.5 g, 861 μmol, 1.0 equiv) in DCM (5.0 mL) was added TFA (1.54 g, 13.5 mmol, 1.0 mL, 15.7 equiv). The mixture was stirred at 20 °C for 12 h. TLC (dichloromethane:methanol = 10:1) showed that the reactants were consumed. The reaction mixture was concentrated under reduced pressure to remove DCM (5.0 mL). The crude product was dissolved in MeOH (50 mL) and adjusted to pH = 7 by adding alkaline resin (NH4HCO3). No purification was performed. Compound 15-6 (0.386 g, crude) was obtained as a yellow oil.

[0247] Step 8. Preparation of compound SIL2-15 [ka] To a solution of compound 15-6 (0.386 g, 935 μmol, 1.0 equiv.) in EtOH (40 mL), 2-decyloxirane (1.38 g, 7.48 mmol, 8.0 equiv.) was added. The mixture was stirred at 120 °C for 72 h. LCMS showed the reactant was consumed, and the product RT = 0.729 min. The reaction mixture was concentrated under reduced pressure to remove EtOH (40 mL). The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 5 / 1 to DCM:MeOH = 30 / 1). Compound SIL2-15 (50.35 mg, 45.0 μmol, 4.82% yield) was obtained as a yellow oil. 1 Checked by 1 H NMR. 1 H NMR: (400 MHz, CDCl) δ ppm 3.58 - 3.70 (m, 4 H), 3.35 - 3.49 (m,5 H), 3.31 - 3.34 (m, 6 H), 2.24 - 3.07 (m, 32 H), 1.16 - 1.56 (m, 75 H), 0.83- 0.94 (m, 12 H).

[0248] Example 9. Synthesis of compound SIL2-17 [ka]

[0249] Step 1. Preparation of compound 17-2 [ka] To a solution of compound 17-1 (2 g, 8.02 mmol, 1 equiv.) in DCM (12 mL) was added EDCI (1.85 g, 9.63 mmol, 1.2 equiv.), TEA (1.62 g, 16.04 mmol, 2.23 mL, 2 equiv.), HOBt (1.30 g, 9.63 mmol, 1.2 equiv.), and compound 17a (1.76 g, 8.82 mmol, 1.1 equiv., HCl). The mixture was stirred at 20 °C for 12 h. TLC (petroleum ether:ethyl acetate = 2:1, R f =0.14), indicating that compound 17-1 was completely consumed and many new spots were formed. The reaction mixture was washed with water (10 mL), the organic phase was separated, and the aqueous phase was extracted with DCM (20 mL × 2), washed with NaHCO (30 mL), brine (30 mL), dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. Compound 17-2 (3.2 g, crude) was obtained as a yellow mixture of a solid and an oil. 1 H NMR: 400 MHz CDCl δ 6.86 (br d, J = 7.88 Hz, 1 H) 5.22 (br d,J = 8.13 Hz, 1 H) 4.70 (td, J = 7.79, 5.07 Hz, 1 H) 4.29 (br d, J = 6.88 Hz, 1H) 3.74 (s, 3 H) 2.58 (t, J = 7.13 Hz, 2 H) 2.50 (t, J = 7.38 Hz, 2 H) 2.12 -2.21 (m, 1 H) 2.09 (d, J = 10.13 Hz, 6 H) 1.88 - 2.06 (m, 3 H) 1.43 (s, 9 H)

[0250] Step 2. Preparation of compound 17-3 [ka] To a solution of compound 17-2 (3.2 g, 8.11 mmol, 1 equiv.) in DCM (8 mL) was added TFA (16.26 g, 142.63 mmol, 10.56 mL, 17.59 equiv.). The mixture was stirred at 20° C. for 12 h. TLC (petroleum ether:ethyl acetate=2:1, Rf = 0.00) indicated that compound 17-2 was completely consumed and one new spot was formed. The reaction mixture was concentrated under reduced pressure to give compound 17-3 (4 g, crude, TFA) as a brown oil.

[0251] Step 3. Preparation of compound 17-4 [ka] Compound 17-3 (4 g, 13.59 mmol, 1 equiv) was dissolved in toluene (40 mL), and TEA (6.87 g, 67.93 mmol, 9.45 mL, 5 equiv) was added, followed by stirring at 130° C. for 12 h. 1 H NMR showed that the starting material was completely consumed. The reaction mixture was diluted with EtOAc (20 mL), filtered, and the filter cake was concentrated. The crude product was used in the next step without further purification. Compound 17-4 (1.7 g, 6.48 mmol, 47.69% yield) was obtained as a white solid. 1 H NMR: 400 MHz DMSO-d6 δ 8.23 ​​(s, 2 H) 3.96 (t, J = 5.50 Hz, 2 H)2.50 - 2.59 (m, 4 H) 2.04 (s, 6 H) 1.83 - 2.02 (m, 4 H)

[0252] Step 4. Preparation of compound C17 [ka] Compound 17-4 (3 g, 11.43 mmol, 1 equiv.) was added portionwise to a solution of LiAlH (1.30 g, 34.30 mmol, 3 equiv.) in THF (30 mL) at 20 °C, and the reaction mixture was stirred at 70 °C for 12 h. LCMS (ET54476-68-P1A1, product: RT = 0.048 min) showed that the starting material was completely consumed. The reaction mixture was cooled to 0 °C and quenched by the dropwise addition of 1.3 mL of water at 0 °C, then diluted with 1.3 mL of 15% NaOH solution and 3.9 mL of water, filtered, and concentrated under reduced pressure to give a residue. Compound C17 (2.9 g, crude) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 2.78 - 2.88 (m, 4 H) 2.68 - 2.75 (m, 2 H)2.52 - 2.60 (m, 4 H) 2.09 - 2.13 (m, 6 H) 1.80 - 1.91 (m, 2 H) 1.68 - 1.77 (m,2 H) 1.48 - 1.64 (m, 2H)

[0253] Step 5. Preparation of compound 17-5 [ka] To a solution of compound 3a (818.22 mg, 3.41 mmol, 1.6 equiv.) in MeCN (25 mL), K2CO3 (884.36 mg, 6.40 mmol, 3 equiv.), NaI (159.85 mg, 1.07 mmol, 0.5 equiv.), and compound C17 (500 mg, 2.13 mmol, 1 equiv.) were added. The mixture was stirred at 90 °C for 12 h. LCMS (ET54476-72-P1A1, product: RT = 0.586 min) showed complete consumption of the starting material. TLC (dichloromethane:methanol = 10:1, R f=0.50) indicated that compound C17 was completely consumed and many new spots were formed. The reaction mixture was filtered, and the filter cake was washed with ACN. It was combined with ET54476-71-p1 for purification. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 20 / 1 to 0 / 1). Compound 17-5 (0.5 g, 30.45% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.97 (br s, 2 H) 3.32 (br d, J = 6.00 Hz,4 H) 2.74 - 2.84 (m, 2 H) 2.52 - 2.72 (m, 16 H) 2.38 - 2.50 (m, 4 H) 2.12 (s, 6H) 1.82 - 1.97 (m, 2 H) 1.69 - 1.80 (m, 2 H) 1.45 (s, 18 H)

[0254] Step 6. Preparation of compound 17-6 [ka] To a solution of compound 17-5 (0.6 g, 936.00 μmol, 1 equiv.) in DCM (0.4 mL) was added TFA (1.08 g, 9.45 mmol, 0.7 mL, 10.10 equiv.). The mixture was stirred at 20° C. for 12 hours. TLC (dichloromethane:methanol=10:1, R f =0.00) indicated that compound 17-5 was completely consumed and one new spot was formed. The reaction mixture was concentrated to remove TFA. The crude product was used in the next step without further purification. Compound 17-6 (0.8 g, crude, TFA) was obtained as a brown oil.

[0255] Step 6. Preparation of compound SIL2-17 [ka] Compound 17-6 (0.4 g, 720.96 μmol, 1 equiv., TFA) was dissolved in methanol, adjusted to pH 8-9 with alkaline resin, filtered, and concentrated to give 180 mg of residue. To a solution of the residue in EtOH (8 mL) was added compound 2a (1.06 g, 5.77 mmol, 8 equiv.). The mixture was stirred at 120 °C for 72 h. LCMS (ET54476-88-P1A2, product: RT = 0.945) showed that the starting material was completely consumed. TLC (dichloromethane:methanol = 10:1, R f =0.34) indicated that compound 17-6 was completely consumed and many new spots were formed. The reaction mixture was concentrated. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 100 / 1 to 0 / 1). Compound SIL2-17 (80 mg, 67.91 μmol, yield 9.42%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 3.63 (br s, 4 H) 3.00 - 3.40 (m, 4 H)2.53 - 2.91 (m, 24 H) 2.27 - 2.51 (m, 10 H) 2.12 (s, 6 H) 1.87 (br d, J = 6.88Hz, 2 H) 1.68 - 1.79 (m, 2 H) 1.24 - 1.48 (m, 72 H) 0.89 (br t, J = 6.75 Hz, 12H) Compounds SIL2-18 to SIL2-21 were prepared in the same manner as above. Their structures and 1H-NMR data are summarized in Table 2. In addition, compounds SIL2-14 and SIL2-16 can also be prepared in the same manner as above. [Table 2] TIFF2025531749000168.tif226149

[0256] Example 10. Synthesis of SIL2-22 [ka]

[0257] Step 1. Preparation of compound 22-2A [ka] To a solution of compound 22-1 (4.00 g, 18.8 mmol, 1.00 equiv) in DCM (24.0 mL) at 0 °C, HOBt (3.04 g, 22.5 mmol, 1.20 equiv) and EDCI (4.32 g, 22.5 mmol, 1.20 equiv) were added. After 30 min, compound B (3.38 g, 20.6 mmol, 1.10 equiv, HCl) and DIEA (3.03 g, 23.5 mmol, 4.08 mL, 1.25 equiv) were added. The reaction was stirred at 25 °C for 7 h. TLC (dichloromethane:methanol = 10:1) showed that the material had been consumed. The DCM solution was washed with 1 N HCl, saturated NaHCO3, and brine, dried over MgSO4, and evaporated in vacuo. Compound 22-2A (6.80 g, crude) was obtained as a brown oil. 1 H NMR: (400 MHz, CDCl) δ ppm 7.12 (d, J=6.00 Hz, 1H), 4.73-4.75(m, 1H), 3.81 (s, 3H), 2.65-2.81 (m, 3H), 2.60-2.65 (m, 1H), 2.12 (d, J=2.40Hz, 1H), 2.02-2.04 (m, 1H), 1.48 (s, 9H).

[0258] Step 2. General Procedure for the Preparation of Compound 22-3A [ka] Compound 22-2A (6.00 g, 19.3 mmol, 1.00 equiv) was dissolved in DCM (10.0 mL), and the mixture was degassed and purged with N2 three times. TFA (11.1 g, 97.0 mmol, 7.20 mL, 5.03 equiv) was added to the mixture at 0 °C. The reaction was stirred at 20 °C for 2 h. TLC (dichloromethane:methanol = 10:1, product Rf =0.10) indicated that the material had been consumed. Concentration under reduced pressure gave a residue. No further purification was carried out. Compound 22-3A (9.20 g, crude) was obtained as a brown oil.

[0259] Step 3. Preparation of compound 22-4A [ka] Compound 22-3A (9.20 g, 43.8 mmol, 1.00 equiv.) was dissolved in Toluene (54.0 mL), degassed, and the mixture was purged with N2 three times. TEA (22.2 g, 219 mmol, 30.5 mL, 5.00 equiv.) was added to the mixture. The mixture was stirred at 120 °C for 12 h. TLC (dichloromethane:methanol = 5:1, material R f = 0.31, dichloromethane:methanol = 20:1, product rf = 0.33) indicated that the material had been consumed. The crude product was concentrated under reduced pressure to give a residue. The crude product was triturated with HO at 20 °C for 30 min. No further purification was carried out. Compound 22-4A (3.00 g, 15.8 mmol, 36.0% yield) was obtained as a white solid. 1 H NMR: (400 MHz, DMSO-d6) δ ppm 8.25 (s, 2H), 4.05 (s, 2H), 2.85 (s,2H), 2.65-2.75 (m, 2H), 2.55 (br s, 2H).

[0260] Step 4. Preparation of compound 22-5 [ka] Compound 22-4A (2.00 g, 11.0 mmol, 1.00 equiv) was dissolved in THF (40.0 mL). LAH (1.40 g, 36.8 mmol, 3.50 equiv) was added to the reaction at 25 °C. The reaction was stirred at 75 °C for 12 h. TLC (dichloromethane:methanol = 5:1, material rf = 0.63) showed material remained. 1.40 mL of HO was added to the reaction at 0 °C, and 1.40 mL of 15% aqueous NaOH and 4.20 mL of HO were poured into the mixture. Anhydrous sodium sulfate was added to the reaction. Filtration and concentration under reduced pressure gave the residue. No further purification was performed. Compound 22-5 (2.75 g, crude) was obtained as a brown solid.

[0261] Step 5. Preparation of compound 22-6 [ka] Compound 22-5 (1.00 g, 6.16 mmol, 1.00 equiv) was dissolved in MeCN (40.0 mL), and K2CO3 (2.56 g, 18.5 mmol, 3.00 equiv) and NaI (462 mg, 3.08 mmol, 0.50 equiv) were added to the mixture at 20 °C. Compound 3a (2.22 g, 9.25 mmol, 1.50 equiv) was added to the mixture at 20 °C. The reaction was stirred at 90 °C for 12 h. TLC (petroleum ether:ethyl acetate = 0:1, product rf = 0.52) indicated that the material had been consumed. Filtration and concentration under reduced pressure gave the residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 2 / 1). Compound 22-6 (0.99 g, 1.74 mmol, 28.2% yield) was obtained as a brown oil. 1 H NMR: (400 MHz, CDCl3) δ ppm 4.97 (s, 1 H), 4.14-4.14 (m, 1 H), 3.32 (s, 4 H), 2.63-2.68 (m, 20 H), 2.01-2.05 (m, 3 H), 1.46 (s, 18 H), 1.27 (t, J=7.20 Hz, 2 H). Step 6.

[0262] Step 6. Preparation of compound 22-7 [ka] Compound 22-6 (0.30 g, 527 μmol, 1.00 equiv) was dissolved in DCM (3.00 mL). TFA (693 mg, 6.08 mmol, 0.45 mL, 11.5 equiv) was added to the mixture at 20 °C. The mixture was stirred for 12 h. TLC (petroleum ether:ethyl acetate = 0:1, material rf = 0.42, product rf = 0.00) showed that the material was consumed. Concentration under reduced pressure gave a residue. Resin (5.00 g) was added to adjust the pH to 7-9. No further purification was performed. Compound 22-7 (0.168 g, 455 μmol, 86.4% yield) was obtained as a brown oil.

[0263] Step 7. Preparation of compound SIL2-22 [ka] Compound 22-7 (0.168 g, 455 μmol, 1.00 equiv) was dissolved in EtOH (40.0 mL). 2-Decyloxirane (672 mg, 3.65 mmol, 8.00 equiv) was added to the mixture. The mixture was stirred at 120 °C for 72 h. TLC (dichloromethane:methanol = 10:1, product rf = 0.49) indicated that the material had been consumed. Concentration under reduced pressure gave a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 0 / 1). Compound SIL2-22 (0.05 g, 28.9 μmol, 6.35% yield) was obtained as a brown oil. 1 H NMR: (400 MHz, CDCl3) δ ppm 6.62 (br s, 1 H), 6.05 (t, J=2.80 Hz,1 H), 5.87 (s, 1 H), 4.02 (t, J=7.20 Hz, 3 H), 3.60 - 3.64 (m, 8 H), 2.98 -3.16 (m, 2 H), 2.26 (s, 18 H), 1.94-1.97 (m, 1 H), 1.21 - 1.27(m, 77 H), 0.89(t, J=6.80 Hz, 12 H). LCMS: (MS / 2+H=553, RT=3.275 min)

[0264] Synthesis of compounds of formula (II) Example 1. Synthesis of compound SIL3-1 Step 1: Preparation of Compound A2-1 [ka] To a solution of compound A (1.50 g, 11.7 mmol, 1.00 equiv.) and compound 1 (1.30 g, 17.6 mmol, 1.61 mL, 1.50 equiv.) in DCM (10 mL) was added DMAP (143 mg, 1.17 mmol, 0.10 equiv.) and DCC (2.66 g, 12.9 mmol, 2.60 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound A remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound A2-1 (2.00 g, 10.9 mmol, 92.7% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.75 - 5.84 (m, 1H), 4.92 - 5.09 (m, 2H),4.08 (t, J = 6.8 Hz, 2H), 2.31 (t, J = 7.6 Hz, 2H), 2.01 - 2.14 (m, 2H), 1.57 -1.69 (m, 5H), 1.31 - 1.50 (m, 4H), 0.94 (t, J = 7.6 Hz, 3H).

[0265] Step 2: Preparation of Compound A2-2 [ka] To a solution of compound A2-1 (2.50 g, 13.6 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (4.39 g, 20.4 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound A2-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (150 mL) and then extracted with petroleum ether (100 mL). The combined organic layers were washed with NaCO (50 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound A2-2 (2.60 g, 13.0 mmol, yield 95.7%) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 4.08 (t, J = 6.8 Hz, 2H), 2.86 - 2.97 (m,1H), 2.76 (t, J = 4.4 Hz, 1H), 2.48 (dd, J = 4.8, 2.8 Hz, 1H), 2.29 - 2.36 (m,2H), 1.62 - 1.74 (m, 3H), 1.46 - 1.60 (m, 5H), 1.33 - 1.44 (m, 2H), 0.94 (t, J= 7.2 Hz, 3H).

[0266] Step 3. Preparation of compound SIL3-1 [ka] To a solution of compound 11 (150 mg, 585 μmol, 1.00 equiv.) in i-PrOH (15 mL) was added compound A2-2 (703 mg, 3.51 mmol, 6.00 equiv.). The mixture was stirred at 120° C. for 12 hours. TLC (dichloromethane:methanol=8:1, product R f =0.31). The mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO, dichloromethane:methanol = 100:1 to 40:1) to give compound SIL3-1 (70.0 mg, 66.2 μmol, yield 11.3%, purity 100%) as a yellow oil. 1 H NMR: 400 MHz CDCl δ 3.08 - 4.08 (m, 13H), 2.86 - 3.07 (m,12H), 2.24 (t, J = 7.3 Hz, 9H), 1.59 - 1.91 (m, 4H), 1.26 - 1.41 (m, 53H), 1.26(br s, 2H), 0.93 (t, J = 7.3 Hz, 12H)

[0267] Example 2. Synthesis of compound SIL3-2 Step 1. Preparation of Compound A4-1 [ka] To a solution of compound A (1.50 g, 11.7 mmol, 1.00 equiv.) and compound 4 (2.04 g, 17.6 mmol, 3.21 mL, 1.50 equiv.) in DCM (10 mL) was added DMAP (1423 mg, 1.17 mmol, 0.10 equiv.) and DCC (2.66 g, 12.9 mmol, 2.60 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f=0.43) indicated that approximately 5% of compound A remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound A4-1 (2.30 g, 10.2 mmol, 86.8% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.75 - 5.86 (m, 1H), 4.94 - 5.04 (m, 2H),4.80 - 4.84 (m, 1H), 2.31 (t, J = 7.6 Hz, 2H), 2.03 - 2.10 (m, 2H), 1.37 - 1.70(m, 8H), 1.20 - 1.35 (m, 4H), 0.80 - 0.95 (m, 6H).

[0268] Step 2. Preparation of Compound A4-2 [ka] To a solution of compound A4-1 (3.00 g, 13.3 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (4.29 g, 19.9 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound A4-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (150 mL) and then extracted with petroleum ether (100 mL). The combined organic layers were washed with NaCO (50 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound A4-2 (3.00 g, 12.4 mmol, yield 93.4%) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 4.82 - 4.88 (m, 1H), 2.87 - 3.02 (m, 1H),2.78 (t, J = 4.8 Hz, 1H), 2.50 (dd, J = 5.2, 2.8 Hz, 1H), 2.35 (t, J = 7.2 Hz,1H), 1.67 - 1.80 (m, 2H), 1.48 - 1.66 (m, 8H), 1.22 - 1.41 (m, 4H), 0.84 - 0.96(m, 6H).

[0269] Step 3. Preparation of compound SIL3-2 [ka] To a solution of compound 11 (100 mg, 390 μmol, 1.00 equiv.) in i-PrOH (10 mL) was added compound A4-2 (567 mg, 2.34 mmol, 6.00 equiv.). The mixture was stirred at 120° C. for 12 hours. TLC (methanol:dichloromethane=1:8) showed one major spot (R f =0.15). The two mixtures were concentrated in vacuo to give the product. The residue was purified by column chromatography (SiO, dichloromethane:methanol = 10 / 1 to 2 / 1). Compound SIL3-2 (50 mg, 40.8 μmol, yield 5.23%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.73 - 4.81 (m, 4H), 3.95 - 4.03 (m, 2H),3.72 - 3.82 (m, 4H), 2.69 - 2.95 (m, 8H), 2.28 - 2.34 (m, 2H), 1.89 - 1.92 (m,5H), 1.20 - 1.67 (m, 70H), 0.82 - 0.91 (m, 24H).

[0270] Example 3. Synthesis of compound SIL3-3 Step 1. Preparation of Compound A5-1 [ka] To a solution of compound 5 (2.04 g, 17.6 mmol, 3.21 mL, 1.50 equiv.) and compound A (1.50 g, 11.7 mmol, 1.00 equiv.) in DCM (10 mL) was added DMAP (143 mg, 1.17 mmol, 0.10 equiv.) and DCC (2.66 g, 12.9 mmol, 2.60 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound A remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound A5-1 (2.20 g, 9.72 mmol, 83.1% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.76 - 5.84 (m, 1H), 4.89 - 5.04 (m, 1H),2.28 (t, J = 7.6 Hz, 2H), 2.01 - 2.06 (m, 2H), 1.53 - 1.70 (m, 3H), 1.38 - 1.52(m, 1H), 1.25 - 1.37 (m, 6H), 1.20 (d, J = 6.0 Hz, 3H), 0.86 - 0.92 (m, 3H).

[0271] Step 2. Preparation of Compound A5-2 [ka] To a solution of compound A5-1 (2.20 g, 9.72 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (3.14 g, 14.58 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound A5-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (150 mL) and then extracted with petroleum ether (100 mL). The combined organic layers were washed with NaCO (50 mL). The combined organic layers were washed with brine (50 mL), dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound A5-2 (2.00 g, 8.25 mmol, 84.9% yield) was obtained as a colorless oil (HNMR: ET40436-15-P1A1). 1 H NMR: 400 MHz CDCl δ 4.86 - 4.95 (m, 1H), 2.86 - 2.97 (m, 1H),2.76 (t, J = 5.2 Hz, 1H), 2.47 (dd, J = 4.8, 2.4 Hz, 1H), 2.30 (t, J = 7.2 Hz,2H), 1.64 - 1.75 (m, 2H), 1.41 - 1.60 (m, 6H), 1.24 - 1.36 (m, 6H), 1.20 (d, J= 6.0 Hz, 3H), 0.81 - 0.95 (m, 3H).

[0272] Step 3. Preparation of compound SIL3-3 [ka] To a solution of compound 11 (150 mg, 585 μmol, 1.00 equiv.) in i-PrOH (15 mL) was added compound A5-2 (851 mg, 3.51 mmol, 6.00 equiv.). The mixture was stirred at 120° C. for 12 hours. TLC (dichloromethane:methanol=10:1) showed that the product R f=0.33). LCMS (ET42086-10-P1A1), product Rt = 0.697 min. The mixture was evaporated to dryness. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 40:1). Compound SIL3-3 (60 mg, 46.45 μmol, yield 3.97%, purity 94.9%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.85 - 4.92 (m, 4H), 4.04 - 4.07 (m, 2H),3.62 - 3.70 (m, 4H), 2.27 - 2.70 (m, 19H), 1.65 - 2.00 (m, 4H), 1.25 - 1.65 (m,63H), 1.20 - 1.28 (m, 12H), 0.87 - 0.90 (m,12H).

[0273] Example 4. Synthesis of compounds SIL3-4 Step 1. Preparation of Compound A6-1 [ka] To a solution of compound A (1.50 g, 11.7 mmol, 1.00 equiv.) and compound 6 (2.29 g, 17.6 mmol, 3.21 mL, 1.50 equiv.) in DCM (10 mL) was added DMAP (143 mg, 1.17 mmol, 0.10 equiv.) and DCC (2.66 g, 12.9 mmol, 2.60 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound A remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound A6-1 (1.20 g, 4.99 mmol, 42.7% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.76 - 5.85 (m, 1H), 4.82 - 5.07 (m, 3H),2.29 (t, J = 7.6 Hz, 2H), 2.06 - 2.12 (m, 2H), 1.61 - 1.69 (m, 2H), 1.38 - 1.52(m, 3H), 1.24 - 1.34 (m, 9H), 1.20 (d, J = 6.4 Hz, 3H), 0.84 - 0.93 (m, 2H),0.84 - 0.93 (m, 1H).

[0274] Step 2. Preparation of Compound A6-2 [ka] To a solution of compound A6-1 (1.90 g, 7.90 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (2.56 g, 11.86 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound A6-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (150 mL) and then extracted with petroleum ether (100 mL). The combined organic layers were washed with NaCO (50 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound A6-2 (2.00 g, 7.80 mmol, yield 98.7%) was obtained as a colorless oil. 1 H NMR: ET40436-16-P1A1). 1 H NMR: 400 MHz CDCl δ 4.86 - 4.95 (m, 1H), 2.88 - 2.95 (m, 1H),2.76 (t, J = 4.8 Hz, 1H), 2.47 (dd, J = 5.2, 2.8 Hz, 1H), 2.26 - 2.35 (m, 2H),1.41 - 1.75 (m, 8H), 1.28 (br s, 7H), 1.20 (d, J = 6.0 Hz, 3H), 0.88 (br t, J =6.4 Hz, 3H).

[0275] Step 3. Preparation of compounds SIL3-4 [ka] To a solution of compound 11 (150 mg, 585 μmol, 1.00 equiv.) in i-PrOH (15 mL) was added compound A6-2 (900 mg, 3.51 mmol, 6.00 equiv.). The mixture was stirred at 120° C. for 12 hours. LCMS (ET42086-5-P1A1, product: R t =0.748 min). TLC (dichloromethane:methanol = 8:1, product R f =0.31). The mixture was concentrated in vacuo. The residue was purified by column chromatography (SiO2, dichloromethane:methanol = 100:1 to 8:1). Compound SIL3-4 (70.0 mg, 52.8 μmol, yield 9.02%, purity 96.7%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 7.27 - 7.73 (m, 2H), 4.86 - 4.93 (m, 4H),4.05 - 4.08 (m, 2H), 3.72 - 3.80 (m, 4H), 2.26 - 2.73 (m, 19H), 2.27 - 2.30 (m,8H), 1.65 - 2.00 (m, 4H), 1.28 - 1.63 (m, 73H), 1.20 - 1.28 (m, 12H), 1.19 -1.20 (m,2H), 0.87 - 0.90 (m, 12H)

[0276] Example 5. Synthesis of Compounds SIL3-5 Step 1. Preparation of Compound A7-1 [ka] To a solution of compound A (1.50 g, 11.7 mmol, 1.00 equiv.) and compound 3 (2.29 g, 17.6 mmol, 3.21 mL, 1.50 equiv.) in DCM (10 mL) was added DMAP (143 mg, 1.17 mmol, 0.10 equiv.) and DCC (2.66 g, 12.9 mmol, 2.60 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound A remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound A7-1 (2.40 g, 9.98 mmol, 85.3% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.75 - 5.84 (m, 1H), 4.91 - 5.10 (m, 2H),3.95 - 4.03 (m, 2H), 2.32 (t, J = 7.6 Hz, 2H), 2.03 - 2.13 (m, 2H), 1.60 - 1.72(m, 2H), 1.52 - 1.57 (m, 1H), 1.22 - 1.49 (m, 10H), 0.82 - 0.96 (m, 6H).

[0277] Step 2. Preparation of Compound A7-2 [ka] To a solution of compound A7-1 (2.40 g, 9.98 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (3.23 g, 14.9 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound A7-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (150 mL) and then extracted with petroleum ether (100 mL). The combined organic layers were washed with NaCO (50 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound A7-2 (2.50 g, 9.75 mmol, 97.7% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 3.89 - 4.06 (m, 2H), 2.88 - 2.97 (m, 1H),2.76 (t, J = 4.4 Hz, 1H), 2.48 (dd, J = 5.2, 2.8 Hz, 1H), 2.30 - 2.39 (m, 2H),1.66 - 1.73 (m, 2H), 1.47 - 1.63 (m, 6H), 1.23 - 1.40 (m, 8H), 0.81 - 0.96 (m,6H).

[0278] Step 3. Preparation of compounds SIL3-5 [ka] A mixture of compound 11 (100 mg, 390 μmol, 1.00 equiv.) and compound A7-2 (600 mg, 2.34 mmol, 6.00 equiv.) in i-PrOH (10 mL) was degassed and purged with N three times, and then the mixture was stirred at 120° C. for 12 h. TLC (methanol:dichloromethane=1:8) confirmed that compound 11 (R f =0.02) was consumed and one major spot (R f=0.25), indicating that no methyl methylcellulose was formed. The two reactions were combined and concentrated in vacuo to give the product. The residue was purified by column chromatography (SiO, dichloromethane:methanol = 1 / 0 to 10 / 1) to give compound SIL3-5 (70.0 mg, 117 μmol, 15.0% yield) as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.03 - 4.13 (m, 2H), 3.90 - 4.01 (m, 7H),3.69 - 3.89 (m, 3H), 2.50 - 2.94 (m, 8H), 2.26 - 2.37 (m, 8H), 1.76 - 2.00 (m,3H), 1.16 - 1.74 (m, 66H), 0.82 - 0.96 (m, 24H)

[0279] Example 6. Synthesis of compound SIL3-12 Step 1. Preparation of Compound B3-1 [ka] To a solution of compound B (3.00 g, 26.3 mmol, 3.12 mL, 1.00 equiv.) and compound 2 (3.47 g, 39.4 mmol, 4.28 mL, 1.50 equiv.) in DCM (20 mL) was added DMAP (321 mg, 2.63 mmol, 0.10 equiv.) and DCC (5.97 g, 28.9 mmol, 5.85 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound B remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound B3-1 (4.50 g, 24.4 mmol, 92.9% yield) was obtained as a colorless oil.

[0280] Step 2. Preparation of Compound B3-2 [ka] To a solution of compound B3-1 (6.00 g, 32.6 mmol, 1.00 equiv.) in DCM (35 mL) was added m-CPBA (10.6 g, 48.8 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound B3-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (300 mL) and then extracted with petroleum ether (200 mL). The combined organic layers were washed with NaCO (100 mL). The combined organic layers were washed with brine (50 mL), dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound B3-2 (4.00 g, 20.0 mmol, yield 61.3%) was obtained as a colorless oil.

[0281] Step 3. Preparation of compound SIL3-12 [ka] A mixture of compound 11 (0.30 g, 1.17 mmol, 1.00 equiv.) and compound B3-2 (1.41 g, 7.02 mmol, 6.00 equiv.) in IPA (30 mL) was degassed and purged with N2 three times, and then the mixture was stirred under N2 atmosphere at 120 °C for 15 h. TLC (dichloromethane:methanol = 10:1, R f=0.17) indicated that the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 30 / 1 to 10 / 1). The residue was purified by preparative HPLC (column: Phenomenex Luna C18 100 × 30 mm × 5 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 40% to 70%, 10 min). The pH was adjusted to 8 with aqueous NaHCO3 and extracted with DCM (10 mL). The combined organic layer was washed with brine (5 mL), filtered, and concentrated under reduced pressure to give a residue. Compound SIL3-12 (0.04 g, 37.83 μmol, yield 3.23%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.47 - 6.71 (m, 2H), 4.06 (t, J = 6.8 Hz,9H), 4.00 - 4.17 (m, 1H), 3.68 (br s, 4H), 2.24 - 2.72 (m, 19H), 1.30 - 1.95(m, 56H), 0.91 (br t, J = 6.4 Hz, 12H).

[0282] Example 7. Synthesis of compounds SIL3-6 Step 1. Preparation of Compound B4-1 [ka] To a mixture of compound B (2.50 g, 21.9 mmol, 2.60 mL, 1.00 equiv.) and compound 4 (3.82 g, 32.9 mmol, 1.50 equiv.) in DCM (25 mL) was added DMAP (268 mg, 2.19 mmol, 0.10 equiv.), followed by DCC (4.97 g, 24.1 mmol, 4.87 mL, 1.10 equiv.) at 0° C. After the addition, the cooling bath was removed and the mixture was stirred at 25° C. for 10 hours. TLC (petroleum ether:ethyl acetate=10:1, R f=0.48), indicating that compound B was completely consumed and two new spots were formed. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO, petroleum ether / ethyl acetate = 50 / 1). Compound B4-1 (3.50 g, 16.5 mmol, 75.3% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.76 - 5.84 (m, 1H), 4.98 - 5.06 (m, 2H),4.80 - 4.87 (m, 1H), 2.32 (t, J = 7.6 Hz, 2H), 2.05 - 2.20 (m, 2H), 1.70 - 1.85(m, 2H), 1.48 - 1.64 (m, 11H), 1.20 - 1.38 (m, 4H), 0.82 - 0.95 (m, 6H).

[0283] Step 2. Preparation of Compound B4-2 [ka] To a solution of compound B4-1 (4.00 g, 18.8 mmol, 1.00 equiv.) in DCM (36 mL) was added m-CPBA (5.28 g, 24.5 mmol, 80% purity, 1.3 equiv.). The mixture was stirred at 25 °C for 10 h. TLC (petroleum ether:ethyl acetate = 10:1) showed that compound B4-1 was completely consumed and many new spots were formed. 1H NMR showed that the desired compound was detected. The mixture was filtered, and the filtrate was diluted with 10% w / v aqueous sodium thiosulfate (200 mL) and stirred vigorously for 5 minutes. The mixture was diluted with saturated aqueous sodium bicarbonate (300 mL), the organic layer was separated, and the aqueous layer was extracted with chloroform (300 mL, 100 mL). The pooled organic extracts were washed with saturated aqueous sodium bicarbonate (100 mL), brine (50 mL), dried over sodium sulfate, and concentrated in vacuo. The crude product was purified by recrystallization from petroleum ether (50 mL) at -10 °C. The mixture was filtered, and the filtrate was concentrated in vacuo. Compound B4-2 (3.20 g, 14.0 mmol, 74.4% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 4.81 - 4.88 (m, 1H), 2.94 - 2.96 (m, 1H),2.78 (t, J = 4.8 Hz, 1H), 2.50 (dd, J = 7.6, 2.8 Hz, 1H), 2.33 - 2.43 (m, 2H),1.75 - 1.93 (m, 2H), 1.47 - 1.69 (m, 6H), 1.21 - 1.41 (m, 4H), 0.80 - 0.98 (m,6H).

[0284] Step 3. Preparation of compounds SIL3-6 [ka] A mixture of compound 11 (0.20 g, 780 μmol, 1.00 equiv.) and compound B4-2 (1.07 g, 4.68 mmol, 6.00 equiv.) in IPA (20 mL) was degassed and purged with N2 three times, and then the mixture was stirred at 120 °C under N2 atmosphere for 15 h. LC-MS showed that the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 40:1 to 15:1, R f =0.17) to give SIL3-6 (0.04 g, 34.20 μmol, 4.38% yield) as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.78 - 4.82 (m, 4H), 3.70 - 4.20 (m, 6H),2.25 - 2.52 (m, 10H), 1.06 - 2.15 (m, 77H), 0.80 - 0.95 (m, 24H).

[0285] Example 8. Synthesis of compounds SIL3-7 Step 1. Preparation of Compound B5-1 [ka] To a mixture of compound B (2.50 g, 21.9 mmol, 2.60 mL, 1.00 equiv.) and compound 5 (3.82 g, 32.8 mmol, 1.50 equiv.) in DCM (25 mL), DMAP (268 mg, 2.19 mmol, 0.10 equiv.) was added, followed by DCC (4.97 g, 24.1 mmol, 4.87 mL, 1.10 equiv.) at 0° C. After the addition, the cooling bath was removed, and the mixture was stirred at 25° C. for 10 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.48), indicating that compound B was completely consumed and two new spots were formed. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO, petroleum ether / ethyl acetate = 50 / 1). Compound B5-1 (4.50 g, 21.2 mmol, yield 96.8%) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.71 - 5.93 (m, 1H), 4.97 - 5.11 (m, 2H),4.83 - 4.96 (m, 1H), 2.30 (t, J = 7.2 Hz, 2H), 2.10 (q, J=7.2 Hz, 2H), 1.68 -1.84 (m, 2H), 1.53 - 1.62 (m, 1H), 1.41 - 1.52 (m, 1H), 1.26 - 1.39 (m, 1H),1.26 - 1.39 (m, 6H), 1.21 (d, J = 6.4 Hz, 3H), 0.89 (br t, J = 6.4 Hz, 3H).

[0286] Step 2. Preparation of Compound B5-2 [ka] To a solution of compound B5-1 (5.00 g, 23.6 mmol, 1.00 equiv.) in DCM (30 mL) was added m-CPBA (6.60 g, 30.6 mmol, 80% purity, 1.30 equiv.). The mixture was stirred at 25 °C for 10 h. TLC (petroleum ether:ethyl acetate = 10:1) showed that compound B5-1 was completely consumed and one new spot was formed. 1 H NMR indicated that the desired compound was detected. The mixture was filtered, and the filtrate was diluted with 10% w / v aqueous sodium thiosulfate (200 mL) and vigorously stirred for 5 min. The mixture was diluted with saturated aqueous sodium bicarbonate (300 mL), the organic layer was separated, and the aqueous layer was extracted with chloroform (300 mL, 100 mL). The pooled organic extracts were washed with saturated aqueous sodium bicarbonate (100 mL), brine (50 mL), dried over sodium sulfate, and concentrated in vacuo. The crude product was purified by recrystallization from petroleum ether (70 mL) at −10 °C. The mixture was filtered, and the filtrate was concentrated in vacuo. Compound B5-2 (5.10 g, 22.3 mmol, 94.8% yield) was obtained as a colorless oil. 1 H NMR:ET40296-15-P1A1 400 MHz CDCl3 δ 4.85 - 5.00 (m, 1H), 2.90 - 2.99 (m, 1H),2.78 (t, J = 4.4 Hz, 1H), 2.50 (dd, J = 5.2, 2.8 Hz, 1H), 2.33 - 2.41 (m, 2H),1.73 - 1.94 (m, 2H), 1.55 - 1.71 (m, 3H), 1.42 - 1.54 (m, 1H), 1.26 - 1.40 (m,6H), 1.22 (d, J = 6.4 Hz, 3H), 0.88 - 0.93 (m, 3H).

[0287] Step 3. General procedure for the preparation of compounds SIL3-7 [ka] A mixture of compound 11 (0.20 g, 780 μmol, 1.00 equiv.) and compound B5-2 (1.07 g, 4.68 mmol, 6.00 equiv.) in IPA (20 mL) was degassed and purged with N2 three times, and then the mixture was stirred at 120 °C under N2 atmosphere for 15 h. LC-MS showed that the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 40:1 to 15:1, R f =0.17) to give compound SIL3-7 (0.05 g, 42.75 μmol, yield 5.48%) as a yellow oil. 1 H NMR: 400 MHz CDCl δ 4.77 - 5.03 (m, 4H), 4.07 (br s, 2H),3.68 - 3.87 (m, 4H), 2.24 - 2.83 (m, 18H), 1.37 - 2.03 (m, 38H), 1.11 - 1.37(m, 38H), 0.86 - 0.92 (m, 12H).

[0288] Example 9. Synthesis of Compound SIL3-8 Step 1. Preparation of Compound B6-1 [ka] To a solution of compound B (3.00 g, 26.3 mmol, 3.12 mL, 1.00 equiv.) and compound 6 (5.13 g, 39.4 mmol, 4.28 mL, 1.50 equiv.) in DCM (20 mL) was added DMAP (321 mg, 2.63 mmol, 0.10 equiv.) and DCC (5.97 g, 28.9 mmol, 5.85 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound B remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound B6-1 (5.50 g, 24.3 mmol, 92.5% yield) was obtained as a colorless oil.

[0289] Step 2. Preparation of Compound B6-2 [ka] To a solution of compound B6-1 (10.0 g, 44.2 mmol, 1.00 equiv.) in DCM (60 mL) was added m-CPBA (14.3 g, 66.3 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 15 h. TLC (petroleum ether:ethyl acetate = 10:1) showed that compound B6-1 was completely consumed and two new spots were formed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (500 mL) and then extracted with petroleum ether (200 mL). The combined organic layers were washed with NaCO (200 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound B6-2 (3.00 g, 12.4 mmol, yield 28.0%) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 4.87 - 4.95 (m, 1H), 2.87 - 2.98 (m, 1H),2.76 (t, J = 4.8 Hz, 1H), 2.48 (dd, J = 4.8, 2.4 Hz, 1H), 2.26 - 2.42 (m, 2H),1.71 - 1.90 (m, 2H), 1.55 - 1.63 (m, 2H), 1.42 - 1.52 (m, 2H), 1.17 - 1.37 (m,11H), 0.89 (t, J = 6.4 Hz, 3H).

[0290] Step 3. Preparation of Compound 6 [ka] A mixture of compound 11 (0.20 g, 780 μmol, 1.00 equiv.) and compound B6-2 (1.13 g, 4.68 mmol, 6.00 equiv.) in IPA (20 mL) was degassed and purged with N2 three times. The mixture was then stirred under N2 atmosphere at 120 °C for 15 h. LC-MS showed that 46.6% of the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 30 / 1 to 10 / 1). The residue was purified by preparative HPLC (column: Phenomenex Luna C18 100 × 30 mm × 5 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 55% to 85%, 10 min). The pH was adjusted to 8 with aqueous NaHCO3 and extracted with DCM (10 mL). The combined organic layers were washed with brine (5 mL), filtered, and concentrated under reduced pressure to give a residue: Compound SIL3-8 (0.045 g, 36.7 μmol, 15.00% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.56 (br s, 1H), 4.84 - 4.95 (m, 4H),4.08 (br s, 2H), 3.74 (br s, 4H), 2.39 - 2.79 (m, 8H), 2.32 (br t, J = 6.8 Hz,8H), 1.38 - 2.05 (m, 34H), 1.16 - 1.35 (m, 50H), 0.86 - 0.91 (m, 12H).

[0291] Example 10. Synthesis of Compound SIL3-9 Step 1. Preparation of compound B7-1 [ka] To a solution of compound B (3.00 g, 26.3 mmol, 3.12 mL, 1.00 equiv.) and compound 3 (5.13 g, 39.4 mmol, 1.50 equiv.) in DCM (20 mL), DMAP (321 mg, 2.63 mmol, 0.10 equiv.) and DCC (5.97 g, 28.9 mmol, 5.85 mL, 1.10 equiv.) were added at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound B remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound B7-1 (5.60 g, 24.7 mmol, 94.1% yield) was obtained as a colorless oil.

[0292] Step 2. Preparation of Compound B7-2 [ka] To a solution of compound B7-1 (7.00 g, 30.9 mmol, 1.00 equiv.) in DCM (45 mL) was added m-CPBA (10.0 g, 46.4 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 25 °C for 12 h. TLC (petroleum ether: ethyl acetate = 10:1) showed that compound B7-1 was completely consumed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (300 mL) and then extracted with petroleum ether (200 mL). The combined organic layers were washed with 100 mL of NaCO. The combined organic layers were washed with 50 mL of brine, dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound B7-2 (4.00 g, 16.5 mmol, 53.4% ​​yield) was obtained as a colorless oil.

[0293] Step 3. Preparation of Compound SIL3-9 [ka] A mixture of compound B (0.30 g, 1.17 mmol, 1.00 equiv.) and compound B7-2 (1.70 g, 7.02 mmol, 6.00 equiv.) in IPA (30 mL) was degassed and purged with N2 three times, and then the mixture was stirred under N2 atmosphere at 120 °C for 15 h. TLC (dichloromethane:methanol = 10:1, R f=0.17) indicated that the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 30 / 1 to 10 / 1). The residue was purified by preparative HPLC (column: Phenomenex Gemini-NX C18 75 × 30 mm × 3 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 50% to 80%, 12 min). The pH was adjusted to 8 with aqueous NaHCO3 and extracted with DCM (10 mL). The combined organic layer was washed with brine (5 mL), filtered, and concentrated under reduced pressure to give a residue. Compound SIL3-9 (62.0 mg, 50.58 μmol, yield 2.16%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.62 (br s, 1H), 3.63 - 4.18 (m, 16H),2.46 - 2.83 (m, 5H), 2.28 - 2.46 (m, 9H), 1.24 - 2.05 (m, 73H), 0.85 - 0.95 (m,24H).

[0294] Example 11. Synthesis of compound SIL3-10 Step 1. Preparation of compound C4-1 [ka] To a solution of compound C (10.0 g, 100 mmol, 10.2 mL, 1.00 equiv.) and compound 4 (17.4 g, 150 mmol, 535 μL, 1.50 equiv.) in DCM (90 mL) was added DMAP (1.22 g, 9.99 mmol, 0.10 equiv.) and DCC (22.7 g, 110 mmol, 22.2 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f=0.43) indicated that approximately 5% of compound C remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound C4-1 (15.0 g, 75.6 mmol, 75.7% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.70 - 5.95 (m, 1H), 4.95 - 5.17 (m, 2H),4.80 - 4.90 (m, 1H), 2.33 - 2.45 (m, 4H), 1.51 - 1.60 (m, 4H), 1.26 - 1.32 (m,5H), 0.86 - 0.91 (m, 7H).

[0295] Step 2. Preparation of compound C4-2 [ka] To a solution of compound C4-1 (15.0 g, 75.6 mmol, 1.00 equiv.) in DCM (100 mL) was added m-CPBA (24.5 g, 113 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 15 °C for 15 h. TLC (petroleum ether:ethyl acetate = 10:1) showed that compound C4-1 was completely consumed and two new spots were formed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (1000 mL) and then extracted with petroleum ether (500 mL). The combined organic layers were washed with NaCO (300 mL). The combined organic layers were washed with brine (100 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1) to give compound C4-2 (5.00 g, 23.3 mmol, yield 30.8%) as a colorless oil. 1 H NMR: 400 MHz CDCl δ 4.80 - 4.87 (m, 1H), 2.93 - 3.04 (m, 1H),2.77 (t, J = 4.8 Hz, 1H), 2.52 (dd, J =4.8, 2.8 Hz, 1H), 2.43 - 2.49 (m, 2H),1.92 - 2.02 (m, 1H), 1.74 - 1.86 (m, 1H), 1.49 - 1.63 (m, 4H), 1.24 - 1.36 (m,4H), 0.86 - 0.91 (m, 6H).

[0296] Step 3. Preparation of compound SIL3-10 [ka] A mixture of compound 11 (0.2 g, 780.20 μmol, 1 equiv.) and compound C4-2 (1.00 g, 4.68 mmol, 6.00 equiv.) in IPA (20 mL) was degassed and purged with N2 three times, and then the mixture was stirred under N2 atmosphere at 120 °C for 15 h. TLC (dichloromethane:methanol = 10:1, R f =0.17) showed that many new spots were formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 30 / 1 to 10 / 1). The residue was purified by preparative HPLC (column: Phenomenex Gemini-NX C18 75 × 30 mm × 3 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 40% to 70%, 10 min). The pH value was adjusted to 8 with aqueous NaHCO3 and extracted with DCM (10 mL). The combined organic layer was washed with 5 mL of brine, filtered, and concentrated under reduced pressure to give a residue. Compound SIL3-10 (114 mg, 102 μmol, yield 38.00%) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.48 (br s, 1H), 4.79 - 4.85 (m, 4H),4.08 (br s, 2H), 3.78 (br s, 4H), 2.29 - 2.89 (m, 20H), 1.42 - 2.11 (m, 38H),1.18 - 1.40 (m, 18H), 0.85 - 0.92 (m, 24H).

[0297] Example 11. Synthesis of compound SIL3-11 Step 1. Preparation of compound C5-1 [ka] To a solution of compound C (15.0 g, 150 mmol, 15.3 mL, 1.00 equiv.) and compound 5 (26.1 g, 225 mmol, 1.50 equiv.) in DCM (90 mL) was added DMAP (1.83 g, 15.0 mmol, 0.10 equiv.) and DCC (34.0 g, 165 mmol, 33.3 mL, 1.10 equiv.) at 0° C. The mixture was stirred at 25° C. for 12 hours. TLC (petroleum ether:ethyl acetate=10:1, R f =0.43) indicated that approximately 5% of compound C remained, and three new major spots were detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 0 to 30 / 1). Compound C5-1 (25.0 g, 126 mmol, 84.1% yield) was obtained as a colorless oil. 1 H NMR: 400 MHz CDCl δ 5.70 - 5.94 (m, 1H), 4.83 - 5.14 (m, 3H),2.32 - 2.44 (m, 4H), 1.44 - 1.64 (m, 2H), 1.23 - 1.37 (m, 6H), 1.20 (d, J = 6.4Hz, 3H), 0.85 - 0.92 (m, 3H).

[0298] Step 2. Preparation of compound C5-2 [ka] To a solution of compound C5-1 (10.0 g, 50.4 mmol, 1.00 equiv.) in DCM (50 mL) was added m-CPBA (16.3 g, 75.6 mmol, 80% purity, 1.50 equiv.) at 0 °C. The mixture was stirred at 15 °C for 15 h. TLC (petroleum ether:ethyl acetate = 10:1) showed that compound C5-1 was completely consumed and two new spots were formed. The reaction mixture was filtered. The filtrate was quenched by adding NaHSO (500 mL) and then extracted with petroleum ether (500 mL). The combined organic layers were washed with NaCO (200 mL). The combined organic layers were washed with brine (50 mL), dried (NaSO), filtered, and concentrated under reduced pressure to give a residue. The crude product was dissolved in petroleum ether. The solid precipitated at -10 °C, filtered, and the filtrate was concentrated under reduced pressure. Compound C5-2 (3.00 g, 14.0 mmol, yield 27.8%) was obtained as a colorless oil.

[0299] Step 3. Preparation of compound C5 [ka] A mixture of compound 11 (0.20 g, 780 μmol, 1.00 equiv.) and compound C5-2 (1.00 g, 4.68 mmol, 6.00 equiv.) in IPA (20 mL) was degassed and purged with N2 three times. The mixture was then stirred under N2 atmosphere at 120 °C for 15 h. LC-MS showed that the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM:MeOH = 30 / 1 to 10 / 1). The residue was purified by preparative HPLC (column: Phenomenex Gemini-NX C18 75 × 30 mm × 3 μm, mobile phase: [water (0.1% TFA)-ACN], B%: 40% to 70%, 10 min). The pH was adjusted to 8 with aqueous NaHCO3 and extracted with DCM (10 mL). The combined organic layers were washed with brine (5 mL), filtered, and concentrated under reduced pressure to give a residue: Compound SIL3-11 (71.0 g, 63.7 μmol, 7.89% yield) was obtained as a yellow oil. 1 H NMR: 400 MHz CDCl δ 6.28 - 6.65 (m, 1H), 4.82 - 4.99 (m, 4H),4.07 (br s, 2H), 3.75 (br s, 3H), 2.22 - 2.81 (m, 18H), 0.95 - 2.07 (m, 72H),0.86 - 0.91 (m, 12H).

[0300] Example 12. Preparation of lipid nanoparticle formulations using a microfluidic device Lipid nanoparticle (LNP) formulations containing lipids of the invention described herein were prepared as follows.

[0301] Briefly, DNA or mRNA solutions were prepared in 10–100 mM citrate buffer at pH 3–5, while the lipid mixture was dissolved in ethanol. The DNA or mRNA aqueous solution and the lipid ethanol solution were mixed in the NanoAssembler at a flow ratio of 5:1–3:1, with a total flow rate of 12 mL / min. The LNP solution was then transferred to a floater and dialyzed against 1x DPBS buffer for 2 hours, then against freshly prepared 1x DPBS for another 2 hours at room temperature. Finally, it was dialyzed against another freshly prepared 1x DPBS solution at 0°C overnight.

[0302] Next, a concentration step was performed. The intermediate LNPs were concentrated 5–15 times using Amicon Ultra-15 (10 kD MWCO) tubes at 2000 g and 23°C. The LNPs were then filtered through a 0.2 μm pore sterile filter. LNP particle size was determined by quasi-elastic light scattering using a Malvern Zetasizer Nano ZS (Malvern, UK), and DNA or mRNA encapsulation and quantification were measured using the Quant-iT RiboGreen mRNA Assay Kit (Thermo Fisher Scientific). Based on the DNA or mRNA quantification results, the LNP solution was further diluted with 1x DPBS buffer and 60 w / w% sucrose solution and stored at -80°C.

[0303] Example 13. Characterization of lipid nanoparticle formulations Once the lipid nanoparticle formulations described in Example 12 were prepared, the size and zeta potential of the lipid nanoparticles, as well as the encapsulation efficiency of nucleic acids into the lipid nanoparticles, were analyzed. Particle size was determined by dynamic light scattering, and zeta potential was measured by electrophoretic light scattering (Zetasizer Nano ZS, Malvern Instruments).

[0304] The encapsulation efficiency (ee%) and quantification of DNA or mRNA into lipid particles were determined using a Ribogreen® (Thermo Scientific) kit. Ribogreen® is an ultrasensitive fluorescent nucleic acid stain for quantifying mRNA, but it can also be used to quantify dsDNA. The ee% was determined by performing a membrane-impermeable fluorescent dye exclusion assay. Briefly, each LNP sample was divided into two groups: one was treated with TE buffer and the other was treated with Triton-X100, a nonionic surfactant known to disrupt LNPs. In addition, a series of DNA or mRNA solutions with known concentrations were also prepared as standard solutions for quantification. All wells were then treated with Ribogreen®, and fluorescence was read after 5–10 minutes. The ee% was calculated as follows: ee% = (I Triton -I TE ) / I Triton is calculated as, where I Triton is the fluorescence of the LNP sample treated with Triton-X100, and I TE was the fluorescence of the LNP group treated with TE buffer. To quantify DNA or mRNA, a standard linear curve was constructed based on the fluorescence intensity and DNA or mRNA standard solutions of known concentrations. To obtain the total quantification of DNA or mRNA in the LNP samples, I Triton Fluorescence values ​​were inserted into the standard linear curve to calculate the DNA or mRNA concentration of the LNP in question.

[0305] Example 14. Cell culture method THP1-dual cell culture Culture flasks (Corning) were removed from the incubator (37°C, 5% CO2, 80% RH). The cell suspension was removed from the culture flask and centrifuged at 1000 rpm for 5 minutes. The supernatant was aspirated, and the cell pellet was reconstituted in growth medium (RPMI (Gibco) containing 10% FBS (Gibco), 1% Pen-Strep (Gibco), and 100 μg / mL Normocin (Invivogen)). An aliquot of the cell suspension was diluted with 0.4% trypan blue (Gibco) for cell counting. Cells were diluted to 5 × 10 in growth medium. 5The cells were transferred to a culture flask at a concentration of 1000 cells / ml, and the flask was placed in an incubator. At each passage, 10 μg / ml of blasticidin (Invivogen) and 100 μg / ml of zeocin (Invivogen) were added to the growth medium.

[0306] To seed cells for the assay, 200 μL / well of cell suspension was added to a 96-well U-bottom tissue culture plate (Costar) at 3 × 10 5 Cells were transferred in cells / mL. Cell seeding was performed the day before the assay.

[0307] HepG2 cell culture Culture flasks (Corning) were removed from the incubator (37°C, 5% CO2, 80% RH). The supernatant was removed from the culture flask, and the cell layer was rinsed with 1x DPBS. Trypsin-EDTA (Gibco) was added to the cell layer, and the flask was placed in the incubator to detach the cells. Growth medium (DMEM (Gibco) containing 10% FBS (Gibco) and 1% Pen-Strep (Gibco)) was added to the flask. An aliquot of the cell suspension was diluted with 0.4% trypan blue (Gibco) to count the cells. The cells were transferred to a gelatin-coated culture flask containing growth medium, and the flask was placed in the incubator.

[0308] To seed cells for the assay, 200 μL of cell suspension was added to a gelatin-coated 96-well flat-bottom tissue culture plate (Costar) at a density of 1.5 × 10 5 Cells were transferred in cells / mL. Cell seeding was performed the day before the assay.

[0309] Example 15. Cell-based assays Gaussia-LNP or hFIX-LNP was diluted to 4 μg / mL in culture medium, and 25 μL was added to each well of a 96-well plate containing 200 μL of THP-1 Dual or HepG2 cells. ApoE4 (Peprotech) was diluted to 10 μg / mL in culture medium, and 25 μL was added to each well of a 96-well plate containing either cell line. The final amount of LNP added and the final ApoE4 concentration in 250 μL of medium were 100 ng and 1 μg / mL per well, respectively.

[0310] THP-1 dual cells were used to detect NFkB and IRF activation 24 hours after LNP stimulation. To detect NFkB activation, the supernatant was collected after 24 hours and diluted 1:10 with Quanti-Blue Solution (InvivoGen). OD values ​​were read at 620 nm absorbance using a microplate reader. To detect IRF activation, the supernatant after 24 hours was diluted 1:3.5 with QUANTI-Luc (InvivoGen). Luminescence units were determined using a luminometer.

[0311] Gaussia expression by hepg2 cells was determined by collecting supernatants 24 hours after LNP stimulation and diluting the supernatants 1:3.5 with QUANIT-Luc. Luminescence units were determined using a luminometer.

[0312] hFIX expression by HepG2 cells was determined by collecting supernatants 24 hours after LNP stimulation and diluting the supernatants 1:1 with assay medium. hFIX concentrations were determined using the FIX ELISA protocol outlined below.

[0313] Example 16. FIX ELISA Assay Protocol A 96-well ELISA plate (Costar) was coated with a capture antibody (Affinity Biologicals), then sealed and incubated at room temperature. The plate was washed three times with PBS-Tween wash buffer and blotted to remove excess liquid. Standards were prepared by serially diluting pooled human normal plasma (George King Biomedical) with mouse control plasma (BioIVT). The prepared standards and samples were diluted with sample diluent. The standard and sample dilutions were transferred to the plate and incubated at room temperature. The plate was washed three times with wash buffer and blotted to remove excess liquid. A detection antibody (Affinity Biologicals) was added to each well of the plate and incubated at room temperature. The plate was washed three times with PBS-Tween wash buffer and blotted to remove excess liquid. TMB (Thermo Scientific) was added to each well of the plate and incubated in the dark at room temperature. Sulfuric acid was added to each well and the plate was read for absorbance at 450 nm on a plate reader (Biotek). A four-parameter curve was constructed against the standards. Sample results were interpolated to the standard curve.

[0314] Example 17. LNP-dsDNA formulations using lipids from the SIL1 series of lipids In one study, LNPs were formulated using the method described in Example 12. These LNPs contained selected lipids of the present invention from the SIL1 series, DSPC, cholesterol, DMG-PEG2000, and erythropoietin (EPO) mRNA. The weight ratio of ionizable lipid to FIX plasmid was between 5 and 15. Table 3 shows exemplary LNPs prepared in this study and their characterization. [Table 3]

[0315] Example 18. In vivo evaluation of LNP1-11 An 11-arm study was conducted in female Balb / C mice for a total duration of 2 days. Eleven groups of mice (6-10 weeks old) weighing approximately 25 grams (n=4 per time point) were administered intravenous (IV) doses as described in Table 4.

[0316] The first plasma sample was collected by serial sampling from Group 1 one day prior to IV dosing. Samples were collected 4 and 24 hours post-dose in all groups. A target total volume of 80 μL of whole blood was collected by retro-orbital bleed. Blood was transferred to lithium heparin tubes and centrifuged at 9,800 × g for 10 minutes at 2–5°C. Plasma was collected and frozen at ≤–70°C until shipped to the sponsor. A total of 40 μL of plasma was the target volume; if less plasma was present, this was noted on the tube. After collection, samples were frozen and stored at ≤–70°C for further analysis. [Table 4]

[0317] Example 19. In vivo study results Plasma hEPO levels were assessed 4 hours later using an EPO-ELISA assay, and the results are shown in Figure 1.

[0318] Example 20. LNP-dsDNA formulations using SIL2 series lipids In one study, LNPs were formulated using the method described in Example 12. These LNPs contained exemplary SIL2 series lipids (SIL2-1 through SIL2-22 and the reference lipid bckk-E12), DOPE, cholesterol, DMG-PEG2000, and C18-PEG2000-TriGalnac ligands (molar ratios of 35:16:46.5:2.4:0.1), and FIX plasmid. The weight ratio of ionizable lipid to FIX plasmid was approximately 10, resulting in an NP ratio of approximately 6.5. Table 5 shows exemplary LNPs prepared in this study and their characterization. [Table 5]

[0319] In conclusion, all LNPs using the novel ionizable SIL-2 lipid were successfully formulated, and all had excellent ee% (>85%). The size of the LNPs ranged from 60 to 75 nm, with a relatively narrow size distribution (PDI <0.2). Consequently, these LNPs were then evaluated in vitro for efficacy and immunogenicity.

[0320] Example 21. In vitro evaluation of LNP915-934 LNP915-934 were tested in vitro using THP-1 cells to assess immunogenicity and in HepG2 cells for efficacy. Cell-based assays were performed according to the protocols outlined in Examples 15 and 16. Briefly, LNP915-934 were incubated with either HepG2 or THP-1 cells at DNA concentrations of either 75 ng or 150 ng per well. hFIX expression by HepG2 cells was determined by collecting supernatants 24 hours after LNP stimulation and diluting the supernatants 1:1 with assay medium (Figures 2 and 3). hFIX concentrations were determined using the FIX ELISA protocol outlined in Example 16. THP-1 dual cells were used to detect NFkB and IRF activation 24 hours after LNP stimulation, and the results are summarized in Figures 4-7.

[0321] Example 22. LNP-mRNA formulations using lipids from the SIL3 series of lipids In one study, LNPs were formulated using the method described in Example 12. These LNPs contained selected ionizable novel SIL3 series lipids, DOPE, cholesterol, and DMG-PEG2000, as well as erythropoietin (EPO) mRNA. The weight ratio of ionizable lipid to FIX plasmid ranged from 5 to 15. Table 6 shows exemplary LNPs prepared in this study and their characterization. [Table 6]

[0322] Example 23. In vivo evaluation of LNP31-42 A 12-arm study was conducted in female Balb / C mice for a total duration of 2 days. Twelve groups of mice (6-10 weeks old) weighing approximately 25 grams (n=4 per time point) were administered intravenous (IV) doses as described in Table 7.

[0323] The first plasma sample was collected by serial sampling from Group 1 one day prior to IV dosing. Samples were collected 4 and 24 hours post-dose in all groups. A target total volume of 80 μL of whole blood was collected by retro-orbital bleed. Blood was transferred to lithium heparin tubes and centrifuged at 9,800 × g for 10 minutes at 2-5°C. Plasma was collected and frozen at ≤-70°C until shipment to the sponsor.

[0324] A total of 40 μL of plasma was the target volume, but if there was less plasma, it was marked as such on the tube. After collection, samples were frozen and stored at ≦−70°C for further analysis. [Table 7]

[0325] Example 24. In vivo test results Plasma hEPO levels were assessed 4 hours later using an EPO-ELISA assay, and the results are shown in FIG.

[0326] Example 25. LNP-DNA formulations using lipids from the SIL3 series of lipids In one study, LNPs were formulated using the method described in Example 12. These LNPs contained selected ionizable novel SIL3 series lipids, DOPE, cholesterol, and DMG-PEG2000, as well as erythropoietin (EPO) mRNA. The weight ratio of ionizable lipid to FIX plasmid ranged from 5 to 15. Table 8 shows exemplary LNPs prepared in this study and their characterization. [Table 8]

[0327] Example 26. In vivo evaluation of LNP43-54 A 12-arm study was conducted in female Balb / C mice for a total duration of 2 days. Twelve groups of mice (6-10 weeks old) weighing approximately 25 grams (n=4 per time point) were administered intravenous (IV) doses as described in Table 9.

[0328] The first plasma sample was collected by serial sampling from Group 1 one day prior to IV dosing. For all groups, samples were collected 4 and 16 hours post-dose on the day of dosing, with additional samples collected at 1 and 2 weeks post-dose. A target total volume of 80 μL of whole blood was collected by retro-orbital bleeding.

[0329] For samples collected before, 4 hours after, and 16 hours after administration, blood was allowed to clot and then centrifuged at 9,800 × g for 10 minutes at 2–5°C to prepare serum, which was frozen at ≤–70°C until shipped to the sponsor.

[0330] For samples collected at 1 and 2 weeks, blood was transferred to lithium heparin tubes and centrifuged at 9,800 × g for 10 minutes at 2–5°C to prepare plasma. Plasma was collected and frozen at ≤–70°C until shipped to the sponsor. A total of 40 μL of plasma was the target volume; if less plasma was present, this was marked on the tube.

[0331] Animal weights were recorded during the study on days -1, 2, 4, week 1, and week 2. After collection, samples were frozen and stored at ≦−70°C. [Table 9]

[0332] Example 27. In vivo study results Plasma hEPO levels were assessed at 1 and 2 weeks using an EPO-ELISA assay and are shown in FIG.

[0333] It will be understood by those skilled in the art that changes may be made to the above-described embodiments without departing from the broad concept of the present invention. It is therefore understood that the present invention is not limited to the particular embodiments disclosed, but is intended to cover modifications within the spirit and scope of the present invention as defined by this description. All documents cited herein are incorporated by reference.

Claims

1. Compounds of formula (I) 【Chemical 1】 or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, n is 0 or 1; X １ and X ２ are different or identical, and each independently represent CHR ３ and R １ and R ２ are different or identical, and C １～４ Alkyl and -C １～４ alkyl-Y, each independently selected from the group consisting of -OR ４ , -SR ４ , or N(R ４ ) ２ and said C １～４ The alkyl group may be selected from halogen and C １～４ optionally substituted with one or more selected from the group consisting of alkyl; R ３ is independently in each occurrence selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, cycloalkyl, and heterocycle, wherein said aryl or cycloalkyl or heterocycle is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, alkoxy, and amino; R ４ is independently in each occurrence alkyl and —C １～４ alkyl-Z, wherein Z is -OR ５ , -SR ５ , or N(R ５ ) ２ wherein said alkyl is optionally substituted with one or more hydroxy groups; R ５ is independently in each occurrence a hydroxyl-substituted alkyl and (—C １～４ alkyl-O) ｍ -C １～４ Alkyl-N(R ６ ) ２ m is 0 or 1; R ６ is a hydroxyl-substituted alkyl; However, R １ and R ２ At least one of １～４ not alkyl].

2. X １ and X ２ The compound of claim 1 , wherein

3. X １ and X ２ is CH ２ 3. The compound according to claim 1 or 2, wherein

4. The compound according to any one of claims 1 to 3, which is a compound of formula (IA). 【Chemistry 2】

5. The compound according to any one of claims 1 to 4, wherein n is 0.

6. R １ Ga-C １～４ The compound of any one of claims 1 to 5, wherein Y is alkyl-Y.

7. R ２ Ga-C １～４ The compound of any one of claims 1 to 6, wherein Y is alkyl-Y.

8. Y is -OR ４ , -SR ４ , or N(R ４ ) ２ , preferably SR ４ The compound according to any one of claims 1 to 7,

9. R ４ Ga-C １～４ The compound of any one of claims 1 to 8, which is alkyl-Z.

10. Z is N (R ５ ) ２ The compound according to any one of claims 1 to 9,

11. R ５ The compound of any one of claims 1 to 10, wherein is a hydroxyl-substituted alkyl.

12. R ５ but, 【Chemistry 3】 and R ７ The compound of any one of claims 1 to 11, wherein is unbranched alkyl.

13. R ５ but, 【Chemistry 4】 The compound according to any one of claims 1 to 12,

14. 【Chemical 5】 【change】 【change】 【change】 The compound according to any one of claims 1 to 13, selected from the group consisting of:

15. R ３ is independently in each occurrence selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy.

16. 16. The compound of claim 15, wherein n is 0.

17. R １ and R ２ 17. The compound of claim 15 or 16, wherein:

18. R １ and R ２ Ga-C １～４ The compound of any one of claims 15 to 17, wherein Y is alkyl-Y.

19. Y is -OR ４ , -SR ４ , or N(R ４ ) ２ , preferably SR ４ The compound according to any one of claims 15 to 18, wherein

20. R ４ Ga-C １～４ The compound of any one of claims 15 to 19, which is alkyl-Z.

21. Z is N (R ５ ) ２ The compound according to any one of claims 15 to 20,

22. R ５ The compound of any one of claims 15 to 21, wherein is a hydroxyl-substituted alkyl.

23. R ５ but, 【Chemistry 6】 and R ７ The compound of any one of claims 15 to 22, wherein is unbranched alkyl.

24. R ５ but, 【Chemistry 7】 The compound according to any one of claims 15 to 23, wherein

25. Compound of formula (I-B1) 【Chemistry 8】 [In the formula, R ３ and R ３’ are different or the same and are each independently selected from the group consisting of alkyl, alkenyl, alkynyl, and cycloalkyl, wherein said alkyl, alkenyl, alkynyl, or cycloalkyl is optionally substituted with one or more selected from the group consisting of hydroxyl, —SH, alkoxyl, alkylthio, aryl, and cycloalkyl, wherein said aryl or cycloalkyl is optionally substituted with one or more selected from the group consisting of halogen, hydroxyl, and alkoxy. The compound according to any one of claims 15 to 24, wherein

26. R ３ and R ３’ 26. The compound of claim 25, wherein:

27. Compound of formula (I-B2) 【Chemistry 9】 27. The compound of claim 26, wherein:

28. [Catalog 10] 【change】 【change】 【change】 【change】 【change】 【change】 【change】 The compound according to any one of claims 15 to 27, selected from the group consisting of:

29. Compound of formula (II) 【Chemistry 11】 or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof [In the formula, p is an integer from 1 to 10; q is an integer from 3 to 8; R ８ is unbranched or branched alkyl.

30. 30. The compound of claim 29, wherein q is 3.

31. 31. The compound of claim 29 or 30, wherein p is 3.

32. 31. The compound of claim 29 or 30, wherein p is 2.

33. 31. The compound of claim 29 or 30, wherein p is 1.

34. [Catalog 12] 【change】 【change】 34. The compound of any one of claims 29 to 33, selected from the group consisting of:

35. A compound according to any one of claims 1 to 34 for use in drug delivery or cancer immunotherapy.

36. 36. The compound of claim 35, wherein the drug is a dsDNA, DNA, mRNA, siRNA, or a small molecule drug.

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