Preparation and application of aminoalcohol lipid molecules

Abstract

The invention relates to the field of chemistry, in particular to preparation and application of aminoalcohol lipid molecules. The invention provides a compound as shown in a formula (I), or a stereoisomer, a pharmaceutically acceptable salt or a prodrug of the compound as shown in the formula (I), and lipid nanoparticles prepared from the compound as shown in the formula (I) are high in lysosome escape capacity, high in delivery efficiency, simple in preparation method and high in yield. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of chemistry, and specifically, to the preparation and application of amino alcohol lipid molecules. Background Art

[0002] The delivery problem is a key issue affecting the performance of nucleic acid drugs. As a promising delivery tool, lipid nanoparticles have shown good delivery effects. Generally, lipid nanoparticles are usually formed by self-assembly of ionizable lipid molecules, auxiliary phospholipids, cholesterol, polyethylene glycol-modified phospholipids, etc., among which ionizable lipid molecules play a crucial role as key components. After being endocytosed by cells, lipid nanoparticles are encapsulated in acidic lysosomes. At this time, the ionizable lipid molecules are protonated, changing from electro-neutral to positively charged, and the ionizable lipids undergo a phase inversion of the membrane, releasing the encapsulated nucleic acid drugs into the cytoplasm, and then exerting their medicinal effects.

[0003] Although lipid nanoparticles have advantages such as low immunogenicity, they still face some key challenges. First, existing lipid molecules generally have relatively low lysosomal escape efficiency and low gene drug delivery efficiency. According to literature research, a large number of lipid nanoparticles are cleared after entering cells, and only 1-2% of lipid nanoparticles can successfully escape from endosomes, severely limiting the efficacy of lipid nanoparticles. In addition, existing ionizable lipid molecules have a risk of biological toxicity, which is the main obstacle affecting their translation from the laboratory to clinical and market. In addition, lipid nanoparticles have poor targeting to non-liver organs, resulting in low drug utilization and off-target risks.

[0004] Therefore, in order to improve the delivery efficiency, safety and targeting of lipid nanoparticles, it is of great significance to develop new ionizable lipid molecules. Summary of the Invention

[0005] The present invention aims to solve at least one of the above technical problems to some extent or at least provide a useful commercial option.

[0006] In a first aspect of the present invention, there is provided a compound having a structure as shown in formula (I), or a stereoisomer, tautomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug of the compound shown in formula (I):

[0007]

[0008] Wherein,

[0009] n is any integer from 1 to 10;

[0010] R 1 、R 2Each independently selected from substituted or unsubstituted C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl, where the substituents of the C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl are each independently C 1 -C 10 hydrocarbyl, ester group or halogen;

[0011] R 3 is selected from -H, -CH 2 CH 2 COO-R 4 、substituted or unsubstituted C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 10 alkynyl, where the substituents of the C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 10 alkynyl are each independently C 1 -C 10 hydrocarbyl, ester group or halogen;

[0012] R 4 is selected from substituted or unsubstituted C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl, where the substituents of the C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl are C 1 -C 10 hydrocarbyl, ester group or halogen.

[0013] The present invention provides a novel ionizable lipid molecule. The amino alcohol lipid compound constructs the skeleton of the lipid molecule with an ester bond commonly found in living organisms. The ester bond can be degraded by lipase in the human body, making the lipid molecule have relatively low toxicity and side effects. The amino alcohol lipid compound contains a long-chain non-polar group and a hydrophilic hydroxyl group, having not only hydrophobic characteristics but also hydrophilic characteristics. The amino alcohol lipid compound can use an unsaturated fatty chain as the hydrophobic end and has a relatively low phase transition temperature.

[0014] According to a specific embodiment of the present invention, n is any integer from 2 to 9.

[0015] According to a specific embodiment of the present invention, the R 1 , R 2 are each independently selected from substituted or unsubstituted C 6 -C 18 alkyl, C 6 -C 18 alkenyl, C 6 -C 18 alkynyl, wherein the substituents of the C 6 -C 18 alkyl, C 6 -C 18 alkenyl, C 6 -C 18 alkynyl are each independently C 1 -C 6 hydrocarbyl, ester group or halogen;

[0016] According to a specific embodiment of the present invention, the R 3 is selected from -H, -CH 2 CH 2 COO-R 4 , substituted or unsubstituted C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, wherein the substituents of the C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl are each independently C 1 -C 6 hydrocarbyl, ester group or halogen;

[0017] According to a specific embodiment of the present invention, the R 4 is selected from substituted or unsubstituted C 6 -C18 Alkyl, C 6 -C 18 Alkenyl, C 6 -C 18 Alkynyl, any one of them, wherein the C 6 -C 18 Alkyl, C 6 -C 18 Alkenyl, C 6 -C 18 Alkynyl substituents are each independently C 1 -C 6 Hydrocarbyl, ester group or halogen.

[0018] According to a specific embodiment of the present invention, n is any integer from 2 to 8.

[0019] According to a specific embodiment of the present invention, the R 1 , R 2 Are each independently selected from substituted or unsubstituted Any one of them, wherein the substituents during substitution are each independently C 1 -C 6 Hydrocarbyl, ester group or halogen.

[0020] According to a specific embodiment of the present invention, the R 3 Is selected from substituted or unsubstituted -H, Wherein the substituents during substitution are each independently C 1 -C 6 Hydrocarbyl, ester group or halogen.

[0021] According to a specific embodiment of the present invention, the R 4 Is selected from substituted or unsubstituted Any one of them, wherein the substituents during substitution are each independently C 1 -C 6 Hydrocarbyl, ester group or halogen.

[0022] According to an embodiment of the present invention, n is any integer from 3 to 6.

[0023] According to an embodiment of the present invention, the R 1 , R 2 Are each independently selected from Any one of them.

[0024] According to an embodiment of the present invention, the R 3 Is selected from -H,

[0025] According to an embodiment of the present invention, the R 4 is selected from any one of them.

[0026] According to an embodiment of the present invention, the compound is selected from any one of the following:

[0027]

[0028]

[0029]

[0030] The second aspect of the present invention provides a method for preparing the compound described in the first aspect, including:

[0031] 1) Reacting the compound represented by formula a with the compound represented by formula b to generate the compound represented by formula (1), wherein the R group in formula b is R 1 or R 2 ;

[0032] 2) Removing the protecting group R 5 from the compound represented by formula (1) to generate the compound represented by formula (2);

[0033] 3) Reacting the compound represented by formula (2) with the compound represented by formula c to generate the compound represented by formula (I),

[0034]

[0035] The preparation method of the amino alcohol lipid compound described in this application is simple, avoiding the introduction of impurities and operation errors during the material transfer and separation and purification processes, and greatly improving the production efficiency.

[0036] According to an embodiment of the present invention, the protecting group R 5 is selected from any one of tert-butoxycarbonyl, benzyloxycarbonyl, p-toluenesulfonyl, and trifluoroacetyl.

[0037] According to an embodiment of the present invention, in step 1), the compound represented by formula a and the compound represented by formula b are reacted under the condition of 70-100 °C.

[0038] According to an embodiment of the present invention, the compound represented by formula a and the compound represented by formula b are reacted under the condition of 80-100 °C, preferably 90 °C.

[0039] According to an embodiment of the present invention, the reaction between the compound represented by formula a and the compound represented by formula b can be carried out under the reaction condition without solvent, or under the reaction condition with DMSO or / and DMF as the solvent.

[0040] According to an embodiment of the present invention, the reaction time of the compound shown in formula a and the compound shown in formula b is 24 - 72 h.

[0041] According to an embodiment of the present invention, the reaction time of the compound shown in formula a and the compound shown in formula b is 32 - 64 h.

[0042] According to an embodiment of the present invention, in step 3), the compound shown in formula (2) and the compound shown in formula c are reacted in an organic solvent, and the organic solvent includes at least one of chloroform, glacial acetic acid, dioxane, and n - hexane.

[0043] According to an embodiment of the present invention, the organic solvent includes a mixture of chloroform and glacial acetic acid, a mixture of dioxane and glacial acetic acid, a mixture of n - hexane and glacial acetic acid, etc.

[0044] According to an embodiment of the present invention, the compound shown in formula (2) and the compound shown in formula c are reacted at 60 - 90 °C, preferably at 75 °C.

[0045] According to an embodiment of the present invention, the reaction time of the compound shown in formula (2) and the compound shown in formula c is 8 - 24 h.

[0046] The third aspect of the present invention provides a method for preparing lipid nanoparticles, and the method includes:

[0047] a) Mixing the compound described in the first aspect with a helper phospholipid, cholesterol, and a PEG - lipid in an organic solvent;

[0048] b) Placing the mixed solution obtained in step a) in a buffer solution in an acidic environment to form lipid nanoparticles.

[0049] Among them, the organic solvent is selected from at least one of methanol, ethanol, propanol, butanol, tetrahydrofuran, and DMSO.

[0050] The lipid nanoparticles prepared by the method of the present invention show good nucleic acid transfection efficiency in both in - vitro and in - vivo experiments.

[0051] According to an embodiment of the present invention, the molar ratio of the compound to the helper phospholipid, cholesterol, and PEG - lipid when mixing is (10 - 70):(5 - 40):(20 - 60):(0.1 - 10).

[0052] According to an embodiment of the present invention, the molar ratio of the compound to the helper phospholipid, cholesterol, and PEG - lipid when mixing is (20 - 60):(10 - 30):(30 - 50):(0.2 - 5).

[0053] According to an embodiment of the present invention, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is (45 - 55):(10 - 20):(35 - 45):(0.2 - 3).

[0054] According to an embodiment of the present invention, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is 50:15:38:1. Optionally, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is 45:10:35:0.2. Optionally, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is 55:20:45:3. Optionally, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is 50:10:38:2. Optionally, the molar ratio of the compound to co - phospholipid, cholesterol, and PEG - lipid when mixed is 50:10:35:1.5.

[0055] According to an embodiment of the present invention, the pH value of the acidic environment is between 3 and 6, preferably a pH value of 4.

[0056] According to an embodiment of the present invention, the buffer solution includes at least one of sodium citrate buffer solution, phosphate buffer solution, phthalic acid buffer solution, ammonium acetate buffer solution, and sodium acetate buffer solution;

[0057] According to an embodiment of the present invention, the co - lipid molecules include at least one of 1,2 - dioleoyl - sn - glycero - 3 - phosphoethanolamine (DOPE) and 1,2 - distearoyl - sn - glycero - 3 - phosphocholine (DSPC).

[0058] The fourth aspect of the present invention provides a lipid nanoparticle, which comprises the compound described in the first aspect or a lipid nanoparticle prepared by the method for preparing a lipid nanoparticle described in the third aspect.

[0059] The fifth aspect of the present invention provides a drug, which comprises the lipid nanoparticle described in the fourth aspect and its content, and the content includes any one of nucleic acid molecules, polypeptides, proteins, and small molecule compounds.

[0060] According to an embodiment of the present invention, the content is nucleic acid, and the drug includes drugs for gene therapy, gene vaccination, antisense therapy, interfering RNA, or nucleic acid transfer.

[0061] According to an embodiment of the present invention, the nucleic acid is RNA or DNA, and the RNA is selected from any one of mRNA, rRNA, miRNA, tRNA, siRNA, and snRNA.

[0062] The sixth aspect of the present invention provides the use of the compound described in the first aspect or the lipid nanoparticle described in the fourth aspect in the preparation of a drug carrier.

[0063] Additional aspects and advantages of the present invention will be given in part in the following description, become apparent in part from the following description, or be learned through the practice of the present invention. Detailed Description of Embodiments

[0064] Embodiments of the present invention will be described in detail below. The embodiments described below are exemplary and are only used to explain the present invention and should not be construed as a limitation of the present invention.

[0065] It should be noted that the terms "first" and "second" are only used for descriptive purposes and should not be construed as indicating or implying relative importance or implicitly specifying the quantity of the indicated technical features. Thus, the features defined with "first" and "second" may explicitly or implicitly include one or more of such features. Further, in the description of the present invention, unless otherwise stated, the meaning of "a plurality" is two or more.

[0066] In this document, the term "comprising" or "including" is an open expression, that is, it includes the content specified in the present invention, but does not exclude other aspects.

[0067] In this document, the terms "optionally", "optional" or "option" generally mean that the subsequent event or condition may but does not necessarily occur, and this description includes the case where the event or condition occurs and the case where the event or condition does not occur.

[0068] Unless otherwise stated, the definitions of groups and terms recorded in the specification and claims of this application, including their definitions as examples, exemplary definitions, preferred definitions, definitions recorded in tables, definitions of specific compounds in embodiments, etc., can be combined and combined with each other arbitrarily. The group definitions and compound structures after such combination and combination should fall within the scope recorded in the specification of this application.

[0069] Unless otherwise defined, all scientific and technical terms used herein have the same meaning as commonly understood by those skilled in the art to which the subject matter of the claims belongs. Unless otherwise stated, all patents, patent applications, and published materials cited throughout this document are incorporated herein by reference in their entirety. If there are multiple definitions for a term in this document, the definitions in this chapter shall prevail.

[0070] Unless otherwise indicated, conventional methods within the skill of the art are employed, such as mass spectrometry, NMR, IR, and UV / Vis spectroscopy and pharmacological methods. Unless otherwise specifically defined, the terms employed herein in connection with descriptions of analytical chemistry, organic synthetic chemistry, and in the context of drugs and pharmaceutical chemistry are known in the art. Standard techniques may be used in chemical synthesis, chemical analysis, drug preparation, formulation and delivery, and the treatment of patients. For example, the instructions of the manufacturer for the kits may be utilized, or the reactions and purifications may be carried out in a manner known in the art or as described in the present application. Generally, the above-mentioned techniques and methods may be implemented according to the descriptions in a number of general and more specific documents cited and discussed in this specification, in accordance with conventional methods well-known in the art. In this specification, groups and their substituents may be selected by those skilled in the art to provide stable structural moieties and compounds. When a substituent is described by a conventional chemical formula written from left to right, the substituent also includes the chemically equivalent substituent obtained when the structural formula is written from right to left. For example, CH 2 O is equivalent to OCH 2 .

[0071] For the numerical ranges recited in the specification and claims of the present application, when the numerical range is understood as "integers", it should be understood that the two endpoints of the range and each integer within the range are recited. For example, "integers from 1 to 6" should be understood as reciting each of 0, 1, 2, 3, 4, 5, and 6. "Integers from 0 to 9" should be understood as reciting each of 0, 1, 2, 3, 4, 5, 6, 7, 8, and 9. When the numerical range is understood as "numbers", it should be understood that the two endpoints of the range, each integer within the range, and each decimal within the range are recited. For example, "numbers from 1 to 10" should be understood as not only reciting each of the integers 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10, but also at least reciting the sum of each of these integers respectively with 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, and 0.9.

[0072] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions, and / or dosage forms that, within the scope of sound medical judgment, are suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0073] The term "pharmaceutically acceptable salt" refers to salts of pharmaceutically acceptable non-toxic acids or bases, including salts of inorganic acids and bases, organic acids and bases. Salts derived from inorganic bases include, but are not limited to, metal salts formed by Al, Ca, Li, Mg, K, Na, and Zn; salts derived from organic bases include, but are not limited to, salts of primary, secondary, or tertiary amines, including naturally occurring substituted or unsubstituted amines, cyclic amines, and basic ion exchange resins, such as ammonium, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, diethanolamine, ethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, caffeine, procaine, choline, betaine, penicillin G, ethylenediamine, glucosamine, methylglucosamine, theobromine, triethanolamine, tromethamine, purine, piperazine, piperidine, N-ethylpiperidine, or polyamine resins; salts derived from inorganic acids and organic acids include, but are not limited to, organic salts formed by sulfuric acid, phosphoric acid, nitric acid, hydrobromic acid, hydrochloric acid, formic acid, acetic acid, propionic acid, benzenesulfonic acid, benzoic acid, phenylacetic acid, salicylic acid, alginic acid, anthranilic acid, camphoric acid, citric acid, ethenesulfonic acid, formic acid, fumaric acid, furoic acid, gluconic acid, glucuronic acid, glutamic acid, glycolic acid, isethionic acid, lactic acid, maleic acid, malic acid, mandelic acid, mucic acid, pamoic acid, pantothenic acid, stearic acid, succinic acid, sulfanilic acid, tartaric acid, p-toluenesulfonic acid, malonic acid, 2-hydroxypropionic acid, oxalic acid, glycolic acid, glucuronic acid, galacturonic acid, citric acid, lysine, arginine, aspartic acid, cinnamic acid, p-toluenesulfonic acid, methanesulfonic acid, ethanesulfonic acid, or trifluoromethanesulfonic acid, etc.

[0074] In addition to pharmaceutically acceptable salts, the present invention also contemplates other salts. They can serve as intermediates in the purification of the compound or in the preparation of other pharmaceutically acceptable salts or can be used for the identification, characterization, or purification of the compounds of the present invention.

[0075] The term "stereoisomer" refers to isomers resulting from different spatial arrangements of atoms in a molecule, including cis-trans isomers, enantiomers, diastereoisomers, and conformational isomers. The stereochemical definitions and conventions used in the present invention generally follow S.P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984) McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S., "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York, 1994.

[0076] The term "protecting group" in organic synthesis refers to a molecule containing two or more functional groups. To protect a particular functional group from being destroyed during a reaction, a reagent is often used to protect it first, and then the protecting agent is removed after the reaction is completed.

[0077] The term "tautomer" refers to a functional group isomer produced by the rapid movement of an atom in a molecule between two positions. The compounds of the present invention may exhibit tautomerism. Tautomers can exist in two or more interconvertible forms. Prototropy tautomers result from the migration of a hydrogen atom covalently bonded between two atoms. Tautomers generally exist in an equilibrium form, and attempting to isolate a single tautomer usually produces a mixture whose physical and chemical properties are consistent with those of a mixture of compounds. The position of the equilibrium depends on the chemical characteristics within the molecule. For example, in many aliphatic aldehydes and ketones such as acetaldehyde, the keto form predominates; while in phenols, the enol form predominates. The present invention encompasses all tautomeric forms of the compounds.

[0078] Some compounds of the present invention may have asymmetric carbon atoms (optical centers) or double bonds. Racemates, diastereoisomers, geometric isomers, and individual isomers are all included within the scope of the present invention.

[0079] The graphical representation of racemates, ambiscalemic and scalemic, or enantiomerically pure compounds herein is from Maehr, J. Chem. Ed. 1985, 62: 114 - 120. Unless otherwise specified, the absolute configuration of a stereocenter is represented by a wedge bond and a dashed bond. When the compounds described herein contain an olefinic double bond or other geometrically asymmetric centers, unless otherwise specified, they include E, Z geometric isomers. Similarly, all tautomeric forms are included within the scope of the present invention.

[0080] The compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)- enantiomers, (R)- and (S)- enantiomers, diastereoisomers, D-isomers, L-isomers, and their racemic mixtures and other mixtures, such as enantiomer- or diastereoisomer-enriched mixtures, all of which mixtures are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers and their mixtures are included within the scope of the present invention.

[0081] The optically active (R)- and (S)-isomers, as well as D- and L-isomers, can be prepared by chiral synthesis, chiral reagents, or other conventional techniques. If an enantiomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, where the resulting mixture of diastereomers is separated and the auxiliary group is cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), it forms a diastereomeric salt with a suitable optically active acid or base, and then the diastereomers are resolved by fractional crystallization or chromatography known in the art, and then the pure enantiomer is recovered. In addition, the separation of enantiomers and diastereomers is usually accomplished by using chromatography with a chiral stationary phase, optionally in combination with chemical derivatization (such as forming a carbamate from an amine).

[0082] The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, including deuterium and variants of hydrogen, provided that the valence of the particular atom is normal and the resulting compound is stable. When the substituent is a keto group (i.e., =O), it means that two hydrogen atoms are replaced. Keto substitution does not occur on an aromatic group. The term "optionally substituted" means that it may or may not be substituted, and unless otherwise specified, the type and number of substituents can be arbitrary based on what is chemically achievable.

[0083] The term "halogen" or "halo" is any one of fluorine, chlorine, bromine, and iodine.

[0084] The term "prodrug" means a compound that can be converted into a biologically active compound of the present invention under physiological conditions or by solvolysis. The prodrugs of the present invention are prepared by modifying the functional groups in the compound, and this modification can be removed by conventional operations or in vivo to obtain the parent compound. Prodrugs include compounds formed by connecting a hydroxyl group or an amino group in the compound of the present invention to any group, and when the prodrug of the compound of the present invention is administered to a mammalian individual, the prodrug is cleaved to form a free hydroxyl group and a free amino group, respectively.

[0085] The compounds of the present invention may contain non-natural proportions of atomic isotopes on one or more atoms constituting the compound. For example, the compound can be labeled with a radioactive isotope, such as deuterium ( 2 H), tritium ( 3 H), iodine-125 ( 125 I), or C-14 ( 14 C). All isotopic compositions of the compounds of the present invention, whether radioactive or not, are included within the scope of the present invention.

[0086] The present invention provides a compound having the structure shown in formula (I), or a stereoisomer, pharmaceutically acceptable salt or prodrug of the compound shown in formula (I):

[0087]

[0088] Wherein,

[0089] n is any integer from 1 to 10;

[0090] R 1 and R 2 are each independently selected from substituted or unsubstituted C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl, wherein the substituents of the C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl are each independently C 1 -C 10 hydrocarbyl, ester group or halogen;

[0091] R 3 is selected from -H, -CH 2 CH 2 COO-R 4 substituted or unsubstituted C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 20 alkynyl, wherein the substituents of the C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 20 alkynyl are each independently C 1 -C 10 hydrocarbyl, ester group or halogen;

[0092] R 4 is selected from substituted or unsubstituted C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl, wherein the C 1-C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 The substituents of alkynyl are C 1 -C 10 hydrocarbyl, ester group or halogen.

[0093] According to the embodiments of the present invention, the compounds provided by the present invention are amino alcohol lipid compounds. The specific structural formula of the preferred amino alcohol lipid compounds is shown in Table 1 as follows:

[0094] Table 1

[0095]

[0096]

[0097] According to the specific embodiments of the present invention, the amino alcohol lipid compounds of the present invention are mixed with co-lipid molecules, cholesterol, and PEG2000-DMG in a certain proportion and dissolved in absolute ethanol; then the mixed solution is quickly injected into a sodium citrate buffer solution, and after stirring, an empty lipid nanoparticle solution can be prepared.

[0098] The inventors of the present application prepared a variety of amino alcohol lipid compounds by using the method for preparing the compounds shown in formula (I) provided by the present invention, and selected the compounds shown as L1-L22 from a variety of amino alcohol lipid compounds to prepare lipid nanoparticles by mixing them with co-lipid molecules, cholesterol, and PEG2000-DMG in a certain proportion. The inventors measured the physicochemical properties of the prepared lipid nanoparticles, and the results showed that the carriers prepared from the amino alcohol lipid compounds of the present invention have sufficient safety and a high encapsulation rate for nucleic acid molecules. Further, the inventors measured the in vitro transfection efficiency of the prepared lipid nanoparticles, and the test samples were mammalian cells. The cell transfection results showed that the fluorescence intensity of the luciferase mRNA-loaded amino alcohol lipid nanoparticles of the present invention was significantly higher than that of the commercially available SM-102 lipid nanoparticles loaded with luciferase mRNA. At the same time, the inventors also measured the in vivo transfection efficiency of the prepared lipid nanoparticles, and the test samples were mammals, preferably mice. The in vivo transfection results showed that the amino alcohol lipid nanoparticles of the present invention have a high encapsulation rate for nucleic acid molecules, can successfully transport nucleic acid molecules into cells and even animals and express them, indicating that the amino alcohol liposomes of the present invention have considerable potential for commercial transformation.

[0099] The present invention provides a compound represented by formula (I), or a stereoisomer, pharmaceutically acceptable salt or prodrug of the compound represented by formula (I), a method for preparing the compound represented by formula (I), its stereoisomer, pharmaceutically acceptable salt or prodrug, and the use of the compound of the present invention in the preparation of drugs.

[0100] Beneficial effects

[0101] (1) The present invention provides a novel ionizable lipid molecule. This amino alcohol lipid compound contains a hydrophobic long chain with a non-polar group and a hydrophilic hydroxyl part, and can effectively encapsulate nucleic acid drugs and deliver them into cells.

[0102] (2) The amino alcohol lipid compound described in the present invention constructs the skeleton of the lipid molecule with an ester bond commonly found in organisms. The ester bond can be degraded by lipase in the human body, making the lipid molecule have less toxic and side effects. This amino alcohol lipid compound contains a long-chain non-polar group and a hydrophilic hydroxyl group, not only having hydrophobic characteristics but also hydrophilic characteristics, and can effectively deliver nucleic acids into the cytoplasm of cells. In particular, because the amino alcohol lipid compound has multiple hydrophobic tails at the same time, its escape ability in lysosomes is increased, and the delivery efficiency of lipid nanoparticles is significantly enhanced.

[0103] (3) This amino alcohol lipid compound can use an unsaturated fatty chain as the hydrophobic end. Since the unsaturated fatty chain has a lower phase transition temperature, it can promote the phase transition of lipid particles in endosomes, destroy the stability of endosomes, and further increase the escape performance of nucleic acid drugs from endosomes.

[0104] (4) The preparation method of the amino alcohol lipid compound described in this application is simple, avoiding the introduction of impurities and operation errors during the material transfer and separation and purification processes, greatly improving the production efficiency. The synthesized amino alcohol lipid compound has good biocompatibility and can achieve safe and efficient intracellular delivery of various drugs.

[0105] There is no particular limitation on the reaction solvent used in each reaction step described in the present invention. Any solvent that can dissolve the starting materials to a certain extent and does not inhibit the reaction is included in the present invention. In addition, many similar modifications, equivalent substitutions, or solvents equivalent to those described in the present invention, solvent combinations, and different ratios of solvent combinations are regarded as within the scope of the present invention.

[0106] The solutions of the present invention will be explained below in conjunction with examples. Those skilled in the art will understand that the following examples are only for illustrating the present invention and should not be regarded as limiting the scope of the present invention. For those not specified in the examples regarding specific techniques or conditions, they shall be carried out according to the techniques or conditions described in the literature in the art or according to the product specifications. For the reagents or instruments not specified regarding the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0107] Unless otherwise specified, the structures of the compounds of the present invention are determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). The unit of NMR shift is 10 -6 (ppm). The solvents for NMR measurement are deuterated dimethyl sulfoxide, deuterated chloroform, deuterated methanol, etc., and the internal standard is tetramethylsilane (TMS).

[0108] The synthesis route of the target compound (I) is shown as follows. It should be noted that all the compounds involved in the present invention are synthesized via the route shown below. Although only the specific preparation processes of compounds L2, L8, L11, and L13 are exemplified in the specification, the preparation principles of the other unexemplified compounds of formula (I) are the same as those of compounds L2, L8, L11, and L13. Therefore, the preparation processes of all the unexemplified compounds of formula (I) should be regarded as within the protection scope of the present invention.

[0109]

[0110] Example 1: Preparation of L2

[0111]

[0112] 3 mmol of N-Boc-1,4-butanediamine and 6 mmol of 1,2-epoxytetradecane were added to a round-bottom flask, and the temperature was raised to 90 °C and stirred for 48 h. The reaction progress was monitored by thin-layer chromatography analysis. After the reaction was completed, the crude product was separated by column chromatography, and the eluent was V 二氯甲烷 :V 甲醇 = 20:1 to obtain the yellow oily product tert-butyl (4-(bis(2-hydroxytetradecyl)amino)butyl)carbamate with a yield of 95.1%.

[0113] 0.8 mmol of tert-butyl (4-(bis(2-hydroxytetradecyl)amino)butyl)carbamate, 8 mL of dichloromethane, and 4 mL of trifluoroacetic acid were added to a 25 mL flask, and the mixture was stirred at 25 °C for 4 h. The reaction progress was monitored by thin-layer chromatography analysis. After the reaction was completed, the crude product was washed to neutral with saturated aqueous sodium carbonate solution, dried over anhydrous sodium sulfate, and the mother liquor was concentrated. The crude product was separated by column chromatography, and the eluent was V 二氯甲烷 :V 甲醇 = 10:1 to obtain the yellow oily 1,1'-((4-aminobutyl)azetidin-1-yl)bis(tetradecan-2-ol) with a yield of 98%.

[0114] Add 0.1 mmol of 1,1'-((4-aminobutyl)azetidinyl)bis(tetradecan-2-ol), 0.1 mmol of tetradecyl acrylate, 10 mL of chloroform, and a drop of glacial acetic acid to a 25 mL flask. Stir under reflux at 75 °C for 15 h, and detect the reaction progress by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 : V 甲醇 = 15:1 to obtain yellow oily L2 with a yield of 83%. Then perform mass spectrometry and NMR analysis. The NMR and mass spectrometry data of L2 are as follows: 1 H NMR (400 MHz, CDCl 3 ) δ 7.42 (s, 1H), 3.85 (s, 2H), 3.40 (d, J = 6.1 Hz, 2H), 2.90 (s, 1H), 2.84 (s, 1H), 2.78–2.69 (m, 2H), 2.64 (d, J = 10.3 Hz, 2H), 1.44 (s, 8H), 1.26 (s, 62H), 0.91–0.83 (m, 9H). ESI-MS (m / z): [M+H] + calcd. for C 49 H 100 N 2 O 4 , 781.8; found, 781.8.

[0115] Example 2: Preparation of L8

[0116]

[0117] Add 3 mmol of N-Boc-3-methyl-1,3-propanediamine and 6 mmol of 1,2-epoxytetradecane to a round-bottom flask, heat to 90 °C and stir for 48 h, and detect the reaction progress by thin-layer chromatography analysis. After the reaction is completed, separate the crude product by column chromatography. The eluent is V 二氯甲烷 : V 甲醇 = 18:1 to obtain yellow oily tert-butyl (3-methyl-(bis(2-hydroxytetradecyl)amino)propyl)carbamate with a yield of 91%.

[0118] Add 0.8 mmol of tert-butyl (3-methyl-(bis(2-hydroxytetradecyl)amino)propyl)carbamate, 8 mL of dichloromethane, and 4 mL of trifluoroacetic acid to a 25 mL flask, stir at 25 °C for 4 h, and detect the reaction progress by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 : V 甲醇= 10:1, to give yellow oily 1,1'-((3-methylaminopropyl)azetidinyl)bis(tetradecan-2-ol) with a yield of 93.5%.

[0119] Add 0.1 mmol of 1,1'-((3-methylaminopropyl)azetidinyl)bis(tetradecan-2-ol), 0.1 mmol of tetradecyl acrylate, 10 mL of chloroform, and a drop of glacial acetic acid to a 25 mL flask. Stir under reflux at 75 °C for 15 h, and monitor the reaction by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 15:1, to give yellow oily L8 with a yield of 81%, and perform mass spectrometry and NMR analysis. NMR and mass spectrometry data of L8: 1 H NMR(400MHz,CDCl 3 )δ7.43(s,1H),3.79(s,2H),3.36(d,J = 6.1Hz,2H),2.90(s,1H),2.84(s,2H),2.75–2.64(m,2H),2.61(d,J = 10.3Hz,2H),1.44(m,4H),1.41(s,8H),1.26(s,64H),0.91–0.83(m,12H).ESI-MS(m / z):[M+H] + calcd.for C 49 H 100 N 2 O 4 ,781.8;found,781.8

[0120] Example 3: Preparation of L11

[0121]

[0122] Add 3 mmol of N-Boc-1,6-hexanediamine and 6 mmol of 1,2-epoxytetradecane to a round-bottom flask, heat to 90 °C and stir for 48 h, and monitor the reaction by thin-layer chromatography analysis. After the reaction is completed, separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 20:1, to give yellow oily tert-butyl (6-(bis(2-hydroxytetradecyl)amino)hexyl)carbamate with a yield of 93.2%.

[0123] Add 0.8 mmol of tert-butyl ((6-(bis(2-hydroxytetradecyl)amino)hexyl)carbamate), 8 mL of dichloromethane, and 4 mL of trifluoroacetic acid to a flask. Stir at 25 °C for 4 h, and monitor the reaction by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 10:1 to obtain yellow oily 1,1'-((6-aminohexyl)azetidinyl)bis(tetradecan-2-ol) with a yield of 97.5%.

[0124] Add 0.1 mmol of 1,1'-((6-aminohexyl)azetidinyl)bis(tetradecan-2-ol), 0.1 mmol of tetradecyl acrylate, 10 mL of chloroform, and a drop of glacial acetic acid to a flask. Reflux and stir at 75 °C for 15 h, and monitor the reaction by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 15:1 to obtain yellow oily L11 with a yield of 81%, and perform mass spectrometry and NMR analysis. The NMR and mass spectrometry data of L11 are as follows: 1 H NMR (400 MHz, CDCl 3 ) δ 7.42 (s, 1H), 3.72 (s, 2H), 3.36 (d, J = 6.1 Hz, 2H), 2.79 (s, 1H), 2.74 (s, 1H), 2.64–2.59 (m, 2H), 2.52 (d, J = 10.3 Hz, 2H), 1.41 (m, 8H), 1.33 (s, 8H), 1.26 (s, 56H), 0.91–0.83 (m, 9H). ESI-MS (m / z): [M+H] + calcd. for C 45 H 92 N 2 O 4 , 725.7; found, 725.7.

[0125] Example 4: Preparation of L13

[0126]

[0127] Add 3 mmol of N-Boc-1,4-butanediamine and 6 mmol of 1,2-epoxytetradecane to a flask. Heat to 90 °C and stir for 48 h, and monitor the reaction by thin-layer chromatography analysis. After the reaction is completed, separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇= 20:1, to obtain yellow oily tert-butyl (4-(bis(2-hydroxytetradecyl)amino)butyl)carbamate with a yield of 95.1%.

[0128] Add 0.8 mmol of tert-butyl (4-(bis(2-hydroxytetradecyl)amino)butyl)carbamate, 8 mL of dichloromethane, and 4 mL of trifluoroacetic acid to a flask, stir at 25 °C for 4 h, and detect the reaction by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate solution until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 10:1, to obtain yellow oily 1,1'-((4-aminobutyl)azetidinyl)bis(tetradecan-2-ol) with a yield of 98%.

[0129] Add 0.1 mmol of 1,1'-((4-aminobutyl)azetidinyl)bis(tetradecan-2-ol), 0.1 mmol of oleyl acrylate, 10 mL of chloroform, and a drop of glacial acetic acid to a 25 mL single-necked flask equipped with a magnetic stirrer, reflux and stir at 75 °C for 15 h, and detect the reaction by thin-layer chromatography analysis. After the reaction is completed, wash the crude product with saturated aqueous sodium carbonate solution until neutral, dry over anhydrous sodium sulfate, concentrate the mother liquor, and separate the crude product by column chromatography. The eluent is V 二氯甲烷 :V 甲醇 = 15:1, to obtain yellow oily L13 with a yield of 83%, and perform mass spectrometry and NMR analysis. NMR and mass spectrometry data of L13: 1 H NMR(400MHz,CDCl 3 )δ7.30(s,1H),5.39 - 5.31(m,2H),3.84(s,2H),3.40(d,J = 6.1Hz,2H),2.92(s,1H),2.81(s,1H),2.68–2.59(m,2H),2.54(d,J = 10.3Hz,2H),1.36(s,8H),1.29(s,76H),0.89–0.83(m,9H).ESI-MS(m / z):[M+H] + calcd.for C 53 H 106 N 2 O 4 ,835.8; found,835.8.

[0130] Example 5: Preparation of lipid nanoparticles

[0131] According to the method described in Example 1, a variety of amino alcohol lipid compounds were prepared. The prepared amino alcohol lipid compounds were mixed and dissolved in absolute ethanol with DOPE (purchased from sigma, product number 850725P), cholesterol, and PEG2000-DMG in a preferred molar ratio of 50:10:38.5:1.5. Then the solution was quickly injected into 3 volumes of sodium citrate buffer solution with a pH of 4, and stirred for 1 minute to prepare the pre-LNP solution.

[0132] Example 6: Determination of the physicochemical properties of lipid nanoparticles

[0133] The particle size, polydispersity index PDI, hemolysis rate, and mRNA loading rate of the lipid nanoparticles in the pre-LNP solution obtained in Example 5 were measured.

[0134] Determination of the particle size and polydispersity index PDI of lipid nanoparticles:

[0135] Using Malvern Zetasizer Nano ZS, the size and polydispersity index PDI of lipid nanoparticles were measured by dynamic light scattering in the 173° backscattering detection mode.

[0136] Determination of the hemolytic properties of lipid nanoparticles:

[0137] Prepare D-PBS buffer solutions with pH 7.4 and 5.5. Dilute the dialyzed pre-LNP to 12.5, 25, 50, 100, 200, and 400 μg / mL respectively. Take fresh mouse blood, centrifuge to remove the supernatant, and wash the blood cells with D-PBS at pH 7.4 several times until the supernatant is colorless and transparent. Then resuspend the red blood cells with buffer solutions at pH 7.4 and 5.5 respectively. According to V 红细胞悬液 :V LNP =1:4, sequentially add cell suspensions with different pH values to LNP solutions with different concentrations and corresponding pH values, mix gently and incubate together. After 4 hours, centrifuge, take the supernatant, and measure the absorbance at a wavelength of 540 nm.

[0138] Hemolysis rate % = [(OD sample - OD negative) / (OD positive – OD negative)] × 100%.

[0139] Determination of the luciferase mRNA cell-level transfection performance of lipid nanoparticles:

[0140] The luciferase mRNA (Fluc mRNA) was dissolved in DEPC aqueous solution, and an equal volume of 50% ethanol solution was added to prepare an ethanol solution with a final concentration of 25%. A pre-LNP solution and the mRNA ethanol solution with a volume ratio of 1.5:1 were mixed. Incubate at 50 °C for 20 minutes to obtain lipid nanoparticles encapsulating mRNA, and dialyze in PBS buffer for 2 hours. The encapsulation efficiency of the lipid nanoparticles was measured using the Quant-it Ribogreen RNA Quantitation Kit (purchased from Thermo, catalog number R11490).

[0141] The results of the physicochemical property determination of the lipid nanoparticles are shown in Table 2. Among them, the measurement results of the particle size and polydispersity index PDI of the lipid nanoparticles show that the particle size of the lipid nanoparticles prepared in Example 5 is distributed in the range of 100 - 200 nm, which can meet the requirements of in vivo delivery; the measurement results of the hemolytic performance of the lipid nanoparticles show that the lipid nanoparticles prepared with the amino alcohol lipid compound of the present invention have a low hemolytic activity (7% - 19%) at pH 7.4, indicating that in the human blood circulation system, the lipid nanoparticles prepared in Example 5 are not likely to cause the rupture and dissolution of human red blood cells and cause harm to the human body, while at pH 5.5, the hemolysis rate is relatively high (70% - 96%), indicating that under acidic conditions, the nucleic acid encapsulated by the lipid nanoparticles prepared in Example 5 is easily released; the measurement results of the luciferase mRNA cell-level transfection performance of the lipid nanoparticles show that the carrier prepared with the amino alcohol lipid compound of the present invention has a high encapsulation efficiency for nucleic acid molecules, can successfully transport the nucleic acid molecules into cells and express them. Among them, the mRNA loading rates of L1 - L22 are relatively high, and basically all can reach more than 90%. The above results indicate that the amino alcohol lipid compound of the present invention is conducive to further development and application.

[0142] Table 2

[0143]

[0144]

[0145]

[0146] The hemolysis rate * represents the ratio at a concentration of 100 ug / ml

[0147] Example 7: Determination of the in vitro transfection efficiency of lipid nanoparticles

[0148] The human bronchial epithelial cell line 16HBE in the logarithmic growth phase was seeded at 25×10 3Inoculate cells at a density of [number of cells / well] into a 48-well cell culture plate. After the cells adhere to the plate, add lipid nanoparticles loaded with mRNA for cell transfection (200 ng mRNA / well), and set up 3 replicates for each sample. After 6 hours of transfection, discard the culture medium, add 50 μL / well of PLB lysis buffer, and lyse the cells for 20 minutes. After centrifuging the cell culture plate, aspirate 10 μL of the lysate into a 96-well black plate, add 50 μL / well of luciferase substrate, and immediately measure the luminescence intensity of the sample using a microplate reader.

[0149] The fluorescence intensity of each group of samples is related to the transfection efficiency of Luc-mRNA into fluorescent protein. As shown in Table 3, using the commercially available ionizable ion lipid SM-102 (purchased from Med Chem Express, catalog number HY-134541) with relatively high transfection efficiency on the market as the control group (100%), it can be seen that after cell transfection, the fluorescence intensity of the luciferase mRNA-loaded amino alcohol lipid nanoparticles described in the present invention is significantly higher than that of the luciferase mRNA-loaded commercial SM-102 lipid nanoparticles. Among them, the average fluorescence intensity of compound L2 can reach up to 5578, indicating that the amino alcohol liposomes described in the present invention have considerable potential for commercial transformation.

[0150] Table 3

[0151]

[0152]

[0153]

[0154] The amino alcohol lipid compounds provided by the present invention use ester bonds commonly found in living organisms to construct the backbone of lipid molecules. The ester bonds can be degraded by lipases in the human body, making the lipid molecules have less toxicity and side effects. The amino alcohol lipid compounds contain long-chain non-polar groups and hydrophilic hydroxyl groups, which not only have hydrophobic characteristics but also hydrophilic characteristics, and can effectively deliver substances into the cytoplasm of cells. In particular, because the amino alcohol lipid compounds have multiple hydrophobic tails at the same time, their escape ability from lysosomes is increased, and the delivery efficiency of lipid nanoparticles is significantly enhanced. It should be noted that the delivery efficiency itself is related to the properties of lipid nanoparticles and has nothing to do with the substances being transported. Therefore, any substance that can be encapsulated in the nanoparticles prepared by the present invention is applicable to the present invention. Here, the substances include but are not limited to nucleic acid molecules, small molecule compounds, polypeptides, proteins, etc.

[0155] The above has described the present invention in detail. The purpose is to enable those skilled in this field to understand the content of the present invention and implement it, and it does not limit the protection scope of the present invention. Any equivalent changes or modifications made according to the spirit and essence of the present invention are covered within the protection scope of the present invention.

[0156] In the description of this specification, the descriptions referring to terms such as "one embodiment", "some embodiments", "examples", "specific examples", "some implementation manners" or "some examples", etc., mean that the specific features, structures, materials or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic expressions of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in a suitable manner in any one or more embodiments or examples. In addition, without contradiction, those skilled in the art can combine and combine the different embodiments or examples described in this specification and the features of different embodiments or examples.

[0157] Although the embodiments of the present disclosure have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting the present disclosure. Those of ordinary skill in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present disclosure.

Claims

1. A compound having the structure shown in formula (I), or a stereoisomer, tautomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug of the compound shown in formula (I): Wherein, n is any integer from 1 to 10; R 1 and R 2 are each independently selected from a substituted or unsubstituted C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl, wherein the substituents of the C 1 -C 20 alkyl, C 2 -C 20 alkenyl, C 2 -C 20 alkynyl are each independently a C 1 -C 10 hydrocarbyl group, an ester group or a halogen; R 3 Selected from -H, -CH 2 CH 2 COO-R 4 , substituted or unsubstituted C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 20 alkynyl, wherein the C 1 -C 10 alkyl, C 2 -C 10 alkenyl, C 2 -C 20 alkynyl substituents are each independently C 1 -C 10 hydrocarbyl, ester group or halogen; R 4 Selected from substituted or unsubstituted C 1 -C 20 -alkyl, C 2 -C 20 -alkenyl, C 2 -C 20 -alkynyl, any one of them, wherein the C 1 -C 20 -alkyl, C 2 -C 20 -alkenyl, C 2 -C 20 -alkynyl substituents are C 1 -C 10 -hydrocarbyl, ester group or halogen.

2. The compound according to claim 1, characterized in that, said n is any integer from 2 to 9; Optionally, said R 1 and R 2 are each independently selected from among substituted or unsubstituted C 6 -C 18 alkyl, C 6 -C 18 alkenyl, C 6 -C 18 alkynyl, wherein the substituents of said C 6 -C 18 alkyl, C 6 -C 18 alkenyl, C 6 -C 18 alkynyl are each independently C 1 -C 6 hydrocarbyl, ester group or halogen; Optionally, said R 3 is selected from -H, -CH 2 CH 2 COO-R 4 , substituted or unsubstituted C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, and any one of them, wherein the substituents of the C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl are each independently C 1 -C 6 hydrocarbyl, ester group or halogen; Optionally, the R 4 is selected from a substituted or unsubstituted C 6 -C 18 alkyl group, a C 6 -C 18 alkenyl group, or a C 6 -C 18 alkynyl group, wherein the substituents of the C 6 -C 18 alkyl group, the C 6 -C 18 alkenyl group, and the C 6 -C 18 alkynyl group are each independently a C 1 -C 6 hydrocarbyl group, an ester group, or a halogen.

3. The compound according to claim 1, characterized in that, said n is any integer from 2 to 8; Optionally, said R 1 , R 2 are each independently selected from any one of substituted or unsubstituted , wherein the substituents in the case of substitution are each independently C 1 -C 6 hydrocarbyl, ester group or halogen; Optionally, said R 3 is selected from substituted or unsubstituted -H, wherein when substituted, the substituents are each independently C 1 -C 6 hydrocarbyl, ester group or halogen; Optionally, said R 4 is selected from substituted or unsubstituted any one of them, wherein when substituted, the substituents are each independently C 1 -C 6 hydrocarbyl, ester group or halogen.

4. The compound according to claim 1, characterized in that, said n is any integer from 3 to 6; Optionally, said R 1 , R 2 are each independently selected from Optionally, said R 3 is selected from -H, Optionally, said R 4 is selected from any one of them.

5. The compound according to claim 1, characterized in that, the compound is selected from any one of the following:

6. A method for preparing the compound according to any one of claims 1-5, characterized in that, comprising: 1) React the compound shown by formula a with the compound shown by formula b to form the compound shown by formula (1), wherein the R group in formula b is R 1 or R 2 ; 2) Remove the protecting group R in the compound of formula (1) 5 to produce the compound of formula (2); 3) Reacting the compound shown in formula (2) with the compound shown in formula c to form the compound shown in formula (I), 7. The method according to claim 6, characterized in that, The protecting group R 5 is selected from any one of tert-butoxycarbonyl, benzyloxycarbonyl, p-toluenesulfonyl, and trifluoroacetyl.

8. A method for preparing lipid nanoparticles, characterized in that, the method comprises: a) Mixing the compound according to any one of claims 1-5 with a helper phospholipid, cholesterol and a PEGylated lipid in an organic solvent; b) Adding the mixed solution obtained in step a) to an acidic buffer solution to obtain lipid nanoparticles, wherein the organic solvent is selected from at least one of methanol, ethanol, propanol, butanol, tetrahydrofuran, DMSO.

9. The method according to claim 8, characterized in that, the molar ratio of the compound to the helper phospholipid, cholesterol, PEGylated lipid during mixing is (10-70):(5-40):(20-60):(0.1-10); Optionally, the molar ratio of the compound to the helper phospholipid, cholesterol, PEGylated lipid during mixing is (20-60):(10-30):(30-50):(0.2-5); Optionally, the pH value of the acidic environment is 3-6; Optionally, the buffer solution includes at least one of sodium citrate buffer solution, phosphate buffer solution, phthalic acid buffer solution, ammonium acetate buffer solution, sodium acetate buffer solution; Optionally, the helper lipid molecule is selected from at least one of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC).

10. A lipid nanoparticle, characterized in that, it contains the compound according to any one of claims 1-5 or the lipid nanoparticle prepared by the method for preparing lipid nanoparticles according to claim 8 or 9.

11. A drug, characterized in that, it includes the lipid nanoparticle according to claim 10 and its content, and the content includes any one of nucleic acid molecules, polypeptides, proteins, small molecule compounds; Optionally, the content is nucleic acid, and the drug includes drugs for gene therapy, gene vaccination, antisense therapy, interfering RNA or nucleic acid transfer; Optionally, the nucleic acid is RNA or DNA, and the RNA is selected from any one of mRNA, rRNA, miRNA, tRNA, siRNA, and snRNA.

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