Single-component spleen-targeted lipid nanoparticle and use thereof

Abstract

Disclosed in the present application are a single-component spleen-targeted lipid nanoparticle and use thereof, and specifically disclosed is use of a compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof in targeting spleen immune cells. An LNP-mRNA complex prepared based on the lipid nanoparticle mediates the delivery of mRNA for stable expression in a spleen, and a prepared LNP-mRNA vaccine for delivering an antigen mRNA has a significant therapeutic effect in a solid tumor model. Compared with a traditional four-component formula, the formula is simpler, and the amount of LNP used is reduced when the LNP delivers a nucleic acid drug, thus improving the safety.

Description

A single-component spleen-targeting lipid nanoparticle and its applications Technical Field

[0001] This application relates to the field of immunotherapy, and more specifically, to LNP-mRNA delivery systems, mRNA, and their uses. Background Technology

[0002] The spleen, as the largest immune organ in humans, plays a crucial role in coordinating adaptive immune responses. This has made it an important target organ for mRNA delivery. Spleen-targeted delivery can enhance immune responses against cancer, promote tolerance to self-antigens and exogenous proteins in autoimmune and allergic diseases, and reduce inflammation in pathogenic inflammation. Lipid nanoparticles, as mRNA delivery carriers, typically accumulate in the liver and lack targeting selectivity for organs outside the liver. Therefore, developing spleen-selective carriers has become an ongoing research direction. Summary of the Invention

[0003] This application first obtained a spleen-targeting lymphocyte (LNP) based on a traditional four-component system, but this LNP still showed partial signal detection in the liver. Based on the traditional four-component system, through formulation optimization and investigation of the active lipid to RNA mass ratio, a single-component spleen-targeting LNP was finally obtained.

[0004] This application provides the following technical solution:

[0005] 1. Use of a compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof in targeting splenic immune cells,

[0006] in:

[0007] G1 and G2 are each independently C2-C24 straight-chain alkylene groups that are optionally substituted with substituents;

[0008] L1 and L2 are each independently -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0009] G3 and G4 are each independently a C2-C24 straight-chain alkylene group, either bonded or optionally substituted with substituents;

[0010] L3 and L4 are each independently a bond, -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0011] G5 and G6 are each independently a C2-C24 straight-chain alkyl group or a C2-C24 straight-chain alkenyl group that is optionally substituted with a substituent.

[0012] Z is either a C1-C12 alkylene group optionally substituted with a substituent or a -R group optionally substituted with a substituent. e G7R f -;

[0013] Among them, R e and R f It is a C1-C12 alkylene group that is optionally substituted with a substituent;

[0014] G7 is -NR g -、-(3-7 membered saturated cycloalkane)-、-(3-7 membered heterocyclic alkane)-、-(3-7 membered cyclic arylene)- or -(3-7 membered cyclic heteroarylene)-;

[0015] R g It is a C1-C12 alkylene group that is optionally substituted with a substituent;

[0016] X and Y are each independently a C1-C12 straight-chain alkyl group optionally substituted with a substituent, or -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring;

[0017] R k R m and R n C1-C12 alkyl groups that may be substituted with substituents;

[0018] The substituents are selected from one or more of deuterium, tritium, halogen atoms, amino, hydroxyl, cyano, nitro or alkylene groups.

[0019] 2. Use of a compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof in the preparation of a medicament targeting splenic immune cells.

[0020] in:

[0021] G1 and G2 are each independently C2-C24 straight-chain alkylene groups that are optionally substituted with substituents;

[0022] L1 and L2 are each independently -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0023] G3 and G4 are each independently a C2-C24 straight-chain alkylene group, either bonded or optionally substituted with substituents;

[0024] L3 and L4 are each independently a bond, -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0025] G5 and G6 are each independently a C2-C24 straight-chain alkyl group or a C2-C24 straight-chain alkenyl group that is optionally substituted with a substituent.

[0026] Z is either a C1-C12 alkylene group optionally substituted with a substituent or a -R group optionally substituted with a substituent. e G7R f -;

[0027] Among them, R e and R f It is a C1-C12 alkylene group that is optionally substituted with a substituent;

[0028] G7 is -NR g -、-(3-7 membered saturated cycloalkane)-、-(3-7 membered heterocyclic alkane)-、-(3-7 membered cyclic arylene)- or -(3-7 membered cyclic heteroarylene)-;

[0029] R g It is a C1-C12 alkylene group that is optionally substituted with a substituent;

[0030] X and Y are each independently a C1-C12 straight-chain alkyl group optionally substituted with a substituent, or -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring;

[0031] R k R m and R n C1-C12 alkyl groups that may be substituted with substituents;

[0032] The substituents are selected from one or more of deuterium, tritium, halogen atoms, amino, hydroxyl, cyano, and nitro groups.

[0033] 3. The use according to item 1 or 2, wherein the compound has the structure shown in formula (IA):

[0034] Among them, G1, G2, G3, G4, G5, G6, L1, L2, L3, L4, X and Y are as defined in item 1 or 2;

[0035] G8 is an optionally substituted C1-C12 alkylene group.

[0036] 4. The use according to item 1 or 2, wherein the compound has the structure shown in formula (IB):

[0037] Among them, G1, G2, G3, G4, G5, G6, G7, L1, L2, L3, L4, R e R f X and Y are as defined in item 1 or 2.

[0038] 5. The use according to item 1 or 2, wherein G1 and G2 are unsubstituted C2-C4 alkylene groups.

[0039] 6. The use as described in item 1 or 2, or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, wherein L1 and L2 are -C(=O)O-.

[0040] 7. The use as described in item 1 or 2, wherein G3 and G4 are keys.

[0041] 8. The use according to item 1 or 2, wherein G3 and G4 are unsubstituted C2-C4 alkylene groups.

[0042] 9. The use as described in item 1 or 2, wherein L3 and L4 are keys.

[0043] 10. The use as described in item 1 or 2, wherein L3 and L4 are -OC(=O)O-.

[0044] 11. The use according to item 1 or 2, wherein G5 and G6 are optionally substituted C4-C16 straight-chain alkenyl groups.

[0045] 12. The use according to item 3, wherein the compound has the structure shown in formula (IA-1):

[0046] Where m is 1 or 3, n is 0 or 1, G5 and G6 are defined as in item 1 or 2, G8 is defined as in item 3, and G9 and G 10 For optionally substituted C1-C5 straight-chain alkanes, -R k -N(R m (R) n - or G9 and G 10 Any one of them, along with the nitrogen it is attached to, forms a ring, R K R is an optionally substituted C1-C5 straight-chain alkylene group. m and R n It is a C1-C5 straight-chain alkane that is optionally substituted.

[0047] 13. The use according to item 3, wherein the compound has the structure shown in formula (IA-2):

[0048] G8 is defined as in item 3, and G5 and G6 are defined as in item 3.

[0049] 14. The use according to item 3, wherein the compound has the structure shown in formula (IA-3):

[0050] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 3.

[0051] 15. The use according to item 3, wherein the compound has the structure shown in formula (IA-4):

[0052] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 3.

[0053] 16. The use according to item 3, wherein the compound has the structure shown in formula (IA-5):

[0054] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 3.

[0055] 17. The use according to item 3, wherein the compound has the structure shown in formula (IA-6):

[0056] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 3.

[0057] 18. The use according to item 4, wherein the compound has the structure shown in formula (IB-1):

[0058] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 4.

[0059] 19. The use according to item 4, wherein the compound has the structure shown in formula (IB-2):

[0060] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 4.

[0061] 20. The use according to item 4, wherein the compound has the structure shown in formula (IB-3):

[0062] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 4.

[0063] 21. The use according to item 4, wherein the compound has the structure shown in formula (IB-4):

[0064] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in item 4.

[0065] 22. The use according to any one of items 12-14, wherein G8 is an optionally substituted C2-C6 alkylene group.

[0066] 23. The use as described in item 12, or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, wherein G9 or G 10 Or both have one of the following structures: methyl, ethyl, 2-hydroxyethyl, -C3H6-N(CH3)(CH3)-.

[0067] 24. The use according to any one of items 12-23, wherein G5 or G6 or both have one of the following structures:

[0068] 25. The use according to any one of items 1-24, wherein the compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof is prepared as a composition for use, preferably,

[0069] The composition further comprises a therapeutic or preventative agent and a carrier for delivering the therapeutic or preventative agent, the carrier comprising a cationic lipid comprising one or more of the compounds represented by formula (I) or pharmaceutically available salts thereof.

[0070] 25. The use according to any one of items 2-24, wherein the drug targeting spleen immune cells is a vaccine or drug for treating or preventing tumors, viral infections or inflammation.

[0071] 26. A composition comprising an active component and a carrier for delivering the active component, the active component being a therapeutic or preventative agent; the carrier being a cationic lipid, the cationic lipid being a compound of formula (I) of any one of items 1 to 25, or a pharmaceutically available salt thereof.

[0072] 27. The composition according to claim 26, wherein the active component is selected from one or more of nucleic acid molecules, small molecule compounds, polypeptides or proteins, preferably, the nucleic acid molecule is selected from one or more of single-stranded DNA, double-stranded DNA, short isomers, agomir, antagomir, antisense molecules, small interfering RNA (siRNA), circular RNA, asymmetric interfering RNA (aiRNA), microRNA (miRNA), dicersubstrate RNA (dsRNA), small hairpin RNA (shRNA), transfer RNA (tRNA), messenger RNA (mRNA), locked nucleic acid (LNA), peptide nucleic acid (PNA) or morpholine ring oligonucleotides.

[0073] 28. The composition according to claim 26, wherein the active component comprises at least one mRNA encoding an antigen or a fragment or epitope thereof, preferably, the mRNA being a monocistronic mRNA or a polycistronic mRNA.

[0074] 29. The composition according to claim 28, wherein the antigen is a tumor-specific antigen or a tumor-associated antigen.

[0075] 30. The composition according to claim 28, wherein the mRNA comprises one or more functional nucleotide analogs selected from one or more of pseudouridine, 1-methyl-pseudouridine, or 5-methylcytosine.

[0076] 31. The compound according to claim 27, wherein the small molecule compound is selected from one or more of the following: antitumor drugs, anti-infective drugs, local anesthetics, antidepressants, anticonvulsants, antibiotics / antibacterial agents, antifungal drugs, antiparasitic drugs, hormones, hormone antagonists, immunomodulators, neurotransmitter antagonists, antiglaucoma agents, anesthetics, or imaging agents.

[0077] 32. The composition according to item 26, wherein the mass ratio of the carrier to the active component is 1:10-50:1, preferably 1.5:1.

[0078] Invention Effects

[0079] The beneficial effects of this invention are:

[0080] 1. The single-component spleen-targeting LNP provided in this application is simpler than the traditional four-component spleen-targeting LNP formulation.

[0081] 2. The single-component spleen-targeting LNP to RNA mass ratio provided in this application is 1.5:1, which is lower than the traditional 10:1, reducing the amount of LNP used when delivering nucleic acid drugs and improving safety.

[0082] 3. The LNP delivery of tumor antigen mRNA method provided in this application has shown good therapeutic effects in solid tumor models.

[0083] 4. The LNP delivery of tumor antigen mRNA provided in this application significantly prolonged the survival of pre-immunized mice in tumor prevention experiments.

[0084] 5. The LNP-delivered tumor antigen mRNA treatment method provided in this application, when used in combination with PD-1, shows better therapeutic effects in solid tumor models. Attached Figure Description

[0085] The accompanying drawings are provided to better understand this application and do not constitute an undue limitation thereof. Wherein:

[0086] Figure 1-1 shows the particle size of the four-component system with different mass ratios of LNP and RNA in Example 14;

[0087] Figures 1-2 show the Zeta potentials of the four-component system with different mass ratios of LNP and RNA in Example 14;

[0088] Figure 2-1 shows the biodistribution of the four-component system of LNP and RNA at different mass ratios in wild-type B / C mice in Example 15 (after dissection);

[0089] Figure 2-2 is a bar chart showing the biodistribution of the four-component system LNP to RNA at different mass ratios in wild-type B / C mice in Example 15 (after dissection);

[0090] Figure 3-1 shows the particle size and zeta potential of different formulations of the two-component system LNP composed of compound 50 and cholesterol in Example 18;

[0091] Figure 3-2 shows the particle size and zeta potential of the two-component system LNP composed of compound 50 and DSPC in Example 18 with different formulations;

[0092] Figure 4-1 shows the particle size of different formulations of the two-component system LNP composed of compound 50 and DOPE in Example 18;

[0093] Figure 4-2 shows the zeta potentials of different formulations of the two-component system LNP composed of compound 50 and DOPE in Example 18;

[0094] Figure 5-1 shows the biodistribution of different formulations of the two-component system LNP composed of compound 50 and DOPE in wild-type B / C mice (after dissection);

[0095] Figure 5-2 is a bar chart showing the biodistribution (after dissection) of different formulations of the two-component system LNP composed of compound 50 and DOPE in wild-type B / C mice in Example 19.

[0096] Figure 6-1 shows the particle size of the single-component system LNP and RNA at different mass ratios in Example 20;

[0097] Figure 6-2 shows the Zeta potentials of the single-component system with different mass ratios of LNP and RNA in Example 20;

[0098] Figure 7 shows the biodistribution of the single-component system LNP to RNA at different mass ratios in wild-type B / C mice in Example 21 (after dissection);

[0099] Figure 8 is a bar chart showing the biodistribution of the single-component system LNP to RNA at different mass ratios in wild-type B / C mice in Example 21 (after dissection);

[0100] Figure 9 shows the biodistribution of the single-component formulations of complexes 53 and 56 in wild-type B / C mice (after dissection);

[0101] Figure 10-1A shows the flow cytometry results of CD86, the activation index of pDC and cDC in the spleen, in Example 23 (statistical results).

[0102] Figure 10-1B shows the statistical results of CD40, the pDC and cDC activation index in the spleen, detected by flow cytometry in Example 23.

[0103] Figure 10-2 shows the percentage of pDCs in the spleen detected by flow cytometry in Example 23 (statistical results);

[0104] Figure 10-3 shows the changes in serum IFNα and IFNy levels (statistical results) in Example 23;

[0105] Figure 11-1 shows the level of IFN-γ secreted by the mouse spleen as detected by Elispot in Example 24;

[0106] Figure 11-2 shows the level of IFN-γ secreted by the mouse spleen as detected by Elispot in Example 24;

[0107] Figure 12-1 shows the statistical results of the flow cytometry analysis of the proportions of various immune cells (B cells, NK cells, macrophages, pDC cells, and cDC cells) in the spleen in Example 24.

[0108] Figure 12-2 shows the flow cytometry results of the proportion of T cells in the spleen in Example 24 (statistical results);

[0109] Figure 13 shows the proportion of CD8+ T cells secreting granulomatous mycotoxin in the spleen in Example 24 (statistical results);

[0110] Figure 14-1 shows the proportion of total IgG in serum in Example 24 (statistical results);

[0111] Figure 14-2 shows the proportion of total IgG1 in serum in Example 24 (statistical results);

[0112] Figure 14-3 shows the proportion of total IgG2c in serum in Example 24 (statistical results);

[0113] Figure 15 shows the tumor size change curves in three groups of mice with B16F10-OVA solid tumors treated with OVA vaccine in Example 25;

[0114] Figure 16 shows the survival curves of three groups of mice with B16F10-OVA solid tumors treated with OVA vaccine in Example 25;

[0115] Figure 17-1 shows the results of mouse body weight and serum ALT detection in Example 25;

[0116] Figure 17-2 shows the results of mouse body weight and serum AST detection in Example 25;

[0117] Figure 17-3 shows the results of mouse body weight and serum ALT and AST detection in Example 25;

[0118] Figure 18-1 shows the flow cytometry results of immune cell phenotypes in peripheral blood in Example 26, including the detection results of immune memory and cytotoxic T cells.

[0119] Figure 18-2 shows the flow cytometry results of immune cell phenotypes in peripheral blood in Example 26, including the detection results of immune memory and cytotoxic T cells.

[0120] Figure 18-3 shows the flow cytometry results of immune cell phenotypes in peripheral blood in Example 26, including the detection results of immune memory and cytotoxic T cells.

[0121] Figure 19 shows the proportions of CD45+ immune cells, Treg cells, and macrophages in the tumor microenvironment as detected by flow cytometry in Example 27 (statistical results).

[0122] Figure 20 shows the M1 / M2 ratio (statistical results) detected by flow cytometry in the tumor microenvironment in Example 27.

[0123] Figure 21-1 shows the flow cytometry detection of pDC activation in the tumor microenvironment in Example 27 (statistical results);

[0124] Figure 21-2 shows the flow cytometry detection of pDC activation in the tumor microenvironment in Example 27 (statistical results);

[0125] Figure 22 shows the tumor size change curves of the three groups of mice in Example 28 when evaluating the preventive effect of the OVA vaccine.

[0126] Figure 23 shows the survival curves of the three groups of mice in Example 28 for evaluating the preventive effect of the OVA vaccine;

[0127] Figure 24 shows the tumor size change curves in three groups of mice with B16F10-OVA solid tumors treated with OVA vaccine combined with PD-1 in Example 29;

[0128] Figure 25 shows the survival curves of three groups of mice with B16F10-OVA solid tumors treated with OVA vaccine combined with PD-1 in Example 29. Detailed Implementation

[0129] the term

[0130] Unless otherwise described, all technical and scientific terms used in this application have the same meaning as commonly understood by one of ordinary skill in the art. For the purposes of interpreting this specification, the following terminology will be applied, and where appropriate, terms used in the singular will also include the plural form, and vice versa. All patents, applications, published applications, and other publications are incorporated herein by reference in their entirety. If any description of terminology set forth herein conflicts with any document incorporated herein by reference, the terminology set forth below shall prevail.

[0131] Unless the context otherwise requires, the word “comprise” and its variations, such as “comprising” and “containing”, are interpreted in an open and inclusive sense, that is, “including, but not limited to”.

[0132] As used herein and unless otherwise stated, the term "lipid" refers to a group of organic compounds, including but not limited to fatty acid esters, and is generally characterized by being poorly soluble in water but soluble in many nonpolar organic solvents. Although lipids are generally weakly water-soluble, certain classes of lipids (e.g., lipids modified with polar groups, such as DMG-PEG2000) have limited water solubility and are soluble in water under certain conditions. Known types of lipids include biomolecules such as fatty acids, waxes, sterols, fat-soluble vitamins, monoglycerides, diglycerides, triglycerides, and phospholipids. Lipids can be classified into at least three categories: (1) "simple lipids," including fats and oils, and waxes; (2) "compound lipids," including phospholipids and glycolipids (e.g., DMG-PEG2000); and (3) "derived lipids," such as steroids. In addition, as used herein, lipids also include lipid-like compounds. The term "lipid-like compound," also simply "lipid," refers to lipid-like compounds (e.g., amphiphilic compounds with lipid-like physical properties).

[0133] As used herein and unless otherwise stated, the term "lipid nanoparticle" or "LNP" refers to a particle having at least one nanometer (nm) scale size (e.g., 1 to 1,000 nm) containing one or more types of lipid molecules. The LNPs provided in this application may further contain at least one non-lipid payload molecule (e.g., one or more nucleic acid molecules) or small molecule drugs. In some embodiments, the LNP comprises a non-lipid payload molecule partially or completely encapsulated within a lipid shell. The payload is a negatively charged molecule (e.g., mRNA encoding a viral protein), and the lipid component of the LNP comprises at least one cationic lipid. Without being bound by theory, it is anticipated that the cationic lipid can interact with the negatively charged payload molecule and promote the incorporation and / or encapsulation of the payload into the LNP during LNP formation. Other lipids that can form a portion of the LNPs provided in this application include, but are not limited to, neutral and charged lipids such as steroids or their analogues, polymer-bound lipids, and various zwitterionic lipids.

[0134] As used herein and unless otherwise stated, the term "cationic lipid" may also be referred to as an active ester, which is ionizable so that it exists in a positively charged or neutral form depending on the pH value. The ionization of cationic lipids affects the surface charge of lipid nanoparticles under different pH conditions. This charge state can affect plasma protein uptake, blood clearance, and tissue distribution (Semple, SC et al., Adv Drug Deliv Rev 32:3-17 (1998)) and the ability to form endosomolytic non-bilayer structures (Hafez, IM et al., Gene Ther 8:1188-1196 (2001)), and is crucial for intracellular delivery of nucleic acids.

[0135] As used in this application and unless otherwise stated, the term "steroid" refers to compounds containing the following carbon skeleton:

[0136] Non-limiting examples of steroids include cholesterol, etc.

[0137] As used herein and unless otherwise stated, the term "neutral lipid" encompasses any lipid molecule present in a neutral or zwitterionic form at or within a selected pH value or range. In some embodiments, the selected useful pH value or range corresponds to the pH conditions of the environment in which the lipid is intended to be used, such as physiological pH. As a non-limiting example, neutral lipids that may be used in conjunction with this disclosure include, but are not limited to, phosphatidylcholines such as 1,2-distearyl-sn-glycerol-3-phosphate choline (DSPC), 1,2-dipalmitoyl-sn-glycerol-3-phosphate choline (DPPC), 1,2-dimyristoyl-sn-glycerol-3-phosphate choline (DMPC), 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphate choline (POPC), and 1,2-dioleoyl-sn-glycerol-3-phosphate choline (DOPC); phosphatidylethanolamines such as 1,2-dioleoyl-sn-glycerol-3-phosphate ethanolamine (DOPE) and 2-((2,3-bis(oleoyloxy)propyl))dimethylammonium)ethyl hydrogen phosphate (DOCP); sphingomyelin (SM); ceramides; and steroids such as sterols, sterols, and their derivatives. The neutral lipids provided in this application may be synthetic or derived from natural sources or compounds (from which they are isolated or modified).

[0138] As used herein and unless otherwise stated, the term "alkyl" refers to a saturated straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms. In one embodiment, the alkyl group has, for example, one to twenty-four carbon atoms (C1-C24 alkyl), four to twenty carbon atoms (C4-C20 alkyl), six to sixteen carbon atoms (C6-C16 alkyl), six to nine carbon atoms (C6-C9 alkyl), one to fifteen carbon atoms (C1-C15 alkyl), one to twelve carbon atoms (C1-C12 alkyl), one to eight carbon atoms (C1-C8 alkyl), or one to six carbon atoms (C1-C6 alkyl) and is connected to the remainder of the molecule by a single bond. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-butyl, n-pentyl, 1,1-dimethylethyl (tert-butyl), 3-methylhexyl, 2-methylhexyl, etc. Unless otherwise stated, alkyl groups are optionally substituted.

[0139] As used herein and unless otherwise stated, the term "alkenyl" refers to a straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing one or more carbon-carbon double bonds. Those skilled in the art will understand that the term "alkenyl" also includes groups having "cis" and "trans" configurations, or "E" and "Z" configurations. In one embodiment, the alkenyl group has, for example, two to twenty-four carbon atoms (C2-C24 alkenyl), four to twenty carbon atoms (C4-C20 alkenyl), six to sixteen carbon atoms (C6-C16 alkenyl), six to nine carbon atoms (C6-C9 alkenyl), two to fifteen carbon atoms (C2-C15 alkenyl), two to twelve carbon atoms (C2-C12 alkenyl), two to eight carbon atoms (C2-C8 alkenyl), or two to six carbon atoms (C2-C6 alkenyl) and is connected to the remainder of the molecule by a single bond. Examples of alkenyl groups include, but are not limited to, vinyl, propenyl, butenyl, pentenyl, pent-1,4-dienyl, etc. Unless otherwise stated, alkenyl groups may optionally be substituted.

[0140] As used herein and unless otherwise stated, the term "alkynyl" refers to a straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing one or more carbon-carbon triple bonds. In one embodiment, the alkynyl group has, for example, two to twenty-four carbon atoms (C2-C24 alkynyl), four to twenty carbon atoms (C4-C20 alkynyl), six to sixteen carbon atoms (C6-C16 alkynyl), six to nine carbon atoms (C6-C9 alkynyl), two to fifteen carbon atoms (C2-C15 alkynyl), two to twelve carbon atoms (C2-C12 alkynyl), two to eight carbon atoms (C2-C8 alkynyl), or two to six carbon atoms (C2-C6 alkynyl) and is connected to the remainder of the molecule by a single bond. Examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, butynyl, pentyynyl, etc. Unless otherwise stated, the alkynyl group is optionally substituted.

[0141] As used herein and unless otherwise stated, the term "cyclization" refers to the partial or complete connection within or between molecules to form a cyclic molecular structure. The connection points may be formed by, but are not limited to, one or more C or N atoms. The cyclic molecular structure may be saturated or unsaturated. Unless otherwise stated, the cyclic molecular portion may optionally be substituted.

[0142] When a group described in this application is referred to as a “substitute,” it may be substituted with one or more of any suitable substituents. Illustrative examples of substituents include, but are not limited to, those found in the exemplary compounds and embodiments provided in this application, as well as: halogen atoms such as F, Cl, Br, or I; cyano; oxo (=O); hydroxyl (-OH); alkyl; alkenyl; alkynyl; cycloalkyl; aryl; -(C=O)OR'; -O(C=O)R'; -C(=O)R'; -OR'; -S(O)xR'; -S-SR'; -C(=O)SR'; -SC (=O)R';-NR'R';-NR'C(=O)R';-C(=O)NR'R';-NR'C(=O)NR'R';-OC(=O)NR'R';-NR'C(=O)OR';-NR'S(O)xNR'R';-NR'S(O)xR';and-S(O)xNR'R', where: R' is independently H, C1-C15 alkyl or cycloalkyl each time it appears, and x is 0, 1 or 2.

[0143] As used herein and unless otherwise stated, the terms “optionally selected” or “optionally” (e.g., optionally substituted) mean that the event or condition described below may or may not occur, and the description includes both the possibility that the event or condition occurs and the possibility that it does not occur. For example, “optionally substituted alkyl” means that the alkyl group may or may not be substituted, and the description includes both substituted alkyl groups and unsubstituted alkyl groups.

[0144] The term "prodrug" refers to a derivative of the compound or nucleic acid drug of this application that, when administered to a patient, can directly or indirectly provide the drug. Prodrugs of the composition can be prepared by modifying functional groups present in the compound or nucleic acid (DNA, ASO, siRNA, mRNA, tRNA) in such a way that the modification can be cleaved in a conventional manner or in vivo to yield the parent compound or nucleic acid. Particularly preferred prodrugs are compounds and nucleic acid drugs that, when administered to a patient, can improve the bioavailability of the composition of this application (e.g., more readily absorbed into the bloodstream), or compounds and nucleic acid drugs that promote the delivery of the parent compound to its site of action (e.g., the lymphatic system). Unless otherwise stated, all prodrug forms of the compounds of this application are within the scope of this application, and various prodrug forms are well known in the art.

[0145] As used in this application and unless otherwise stated, the term "pharmaceutically acceptable salt" includes both acid addition salts and base addition salts.

[0146] Examples of pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid; and organic acids, such as, but not limited to, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, 4-acetamidobenzoic acid, camphoric acid, camphor-10-sulfonic acid, decanoic acid, hexanoic acid, caprylic acid, carbonic acid, cinnamic acid, citric acid, and cyclamic acid. (acid), dodecyl sulfate, ethane-1,2-disulfonic acid, ethane sulfonic acid, 2-hydroxyethane sulfonic acid, formic acid, fumaric acid, galactosic acid, gentian acid, glucoheponic acid, gluconic acid, glucuronic acid, glutamic acid, glutamate, 2-oxoglutamate, glycerophosphate, glycolic acid, hippuric acid, isobutyric acid, lactic acid, lactobionic acid, lauric acid, maleic acid, malic acid, malonic acid, mandelic acid, methane sulfonic acid, mucoic acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid, oleic acid, orotic acid, oxalic acid, palmitic acid, papoic acid, propionic acid, pyroglutamic acid, pyruvic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, tartaric acid, thiocyanate, p-toluenesulfonic acid, trifluoroacetic acid, undecenoic acid, etc.

[0147] Examples of pharmaceutically acceptable base addition salts include, but are not limited to, salts prepared by adding an inorganic or organic base to a free acid compound. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. In one embodiment, the inorganic salt is an ammonium salt, sodium salt, potassium salt, calcium salt, or magnesium salt. Salts derived from organic bases include, but are not limited to, the following: primary, secondary, and tertiary amines; substituted amines, including naturally occurring substituted amines; cyclic amines and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, diethanolamine, ethanolamine, deanol, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, benethamine, benzathine, ethylenediamine, glucosamine, methylglucosamine, theobromine, triethanolamine, aminobutanetriol, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc.

[0148] The compounds provided in this application may contain one or more asymmetric centers, and thus may produce enantiomers, diastereomers, and other stereoisomers, which may be defined by absolute stereochemistry as (R)- or (S)-, or for amino acids as (D)- or (L)-. Unless otherwise stated, the compounds provided in this application are intended to include all such possible isomers, as well as their racemic and optically pure forms. When the compounds of this application contain an alkene double bond or other geometrically asymmetric centers, the compounds are intended to include E and Z geometric isomers, unless otherwise stated. Similarly, all tautomers are also intended to be included.

[0149] As used herein and unless otherwise stated, the term "isomer" refers to different compounds having the same molecular formula. "Stereoisomer" is an isomer that differs only in the spatial arrangement of its atoms. "Restricted rotational isomer" is a stereoisomer obtained by restricted rotation around a single bond. "Enantiomer" is a pair of stereoisomers that are non-overlapping mirror images of each other. A mixture of any proportion of a pair of enantiomers may be called a "racemic" mixture. "Diadiaomer" is a stereoisomer having at least two asymmetric atoms but not being mirror images of each other.

[0150] "Stereoisomers" may also include E and Z isomers or mixtures thereof, as well as cis and trans isomers or mixtures thereof. In some embodiments, the compound described in this application is isolated as an E or Z isomer. In other embodiments, the compound described in this application is a mixture of E and Z isomers.

[0151] The term "nucleic acid" refers to a polymer of nucleotides of any length and includes, for example, DNA and RNA. Nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases and / or their analogues, or any substrate that can be incorporated into the polymer by DNA or RNA polymerase or by a synthetic reaction. Nucleic acids can be single-stranded or double-stranded. As used herein and unless otherwise stated, "nucleic acid" also includes nucleic acid mimics such as locked nucleic acids (LNA), peptide nucleic acids (PNA), and morpholine nucleic acids. As used herein, "oligonucleotide" refers to a short synthetic polynucleotide whose length is generally, but not necessarily, less than about 200 nucleotides. The terms "oligonucleotide" and "polynucleotide" are not mutually exclusive. The above description of polynucleotides applies equally and fully to oligonucleotides. Unless otherwise stated, the left-hand end of any single-stranded polynucleotide sequence disclosed herein is the 5' end; the left-hand direction of a double-stranded polynucleotide sequence is referred to as the 5' direction. The direction of addition from 5′ to 3′ to the nascent RNA transcript is called the transcription direction; the region on the DNA strand that has the same sequence as the RNA transcript and is located at the 5′ end relative to the 5′ end of the RNA transcript is called the “upstream sequence”; the region on the DNA strand that has the same sequence as the RNA transcript and is located at the 3′ end relative to the 3′ end of the RNA transcript is called the “downstream sequence”.

[0152] "Isolated nucleic acid" refers to nucleic acid, such as RNA, DNA, or a mixture of nucleic acids, that is substantially isolated from other genomic DNA sequences naturally accompanying their natural sequences, as well as proteins or complexes (such as ribosomes and polymerases). An "isolated" nucleic acid molecule is a nucleic acid molecule isolated from other nucleic acid molecules present in the natural source of the nucleic acid molecule. Furthermore, when manufactured using recombinant technology, an "isolated" nucleic acid molecule, such as an mRNA molecule, may be substantially free of other cellular material or culture medium, or when chemically synthesized, it may be substantially free of chemical precursors or other chemicals. In a particular embodiment, one or more nucleic acid molecules encoding antigens described in this application are isolated or purified. The term includes nucleic acid sequences that have been removed from their natural environment and includes recombinant or cloned DNA or RNA isolates, as well as chemically synthesized analogs or analogs biosynthesized from heterologous systems. A substantially pure molecule may include an isolated form of the molecule.

[0153] The term “coding nucleic acid” or its grammatical equivalent, when used to refer to nucleic acid molecules, includes: (a) nucleic acid molecules that, in their natural state or by methods well known to those skilled in the art, can be transcribed to produce mRNA and subsequently translated into peptides and / or polypeptides; and (b) the mRNA molecule itself. An antisense sequence is a complementary sequence of such nucleic acid molecules from which the coding sequence can be inferred. The term “coding region” refers to the portion of a nucleic acid sequence that is translated into a peptide or polypeptide. The term “untranslated region” or “UTR” refers to the portion of a nucleic acid sequence that is not translated into a peptide or polypeptide. Depending on the orientation of the UTR relative to the coding region of the nucleic acid molecule, a UTR located at the 5′ end of the coding region is called a 5′-UTR, and a UTR located at the 3′ end of the coding region is called a 3′-UTR.

[0154] As used herein, the term "mRNA" refers to a messenger RNA molecule containing one or more open reading frames (ORFs) that can be translated by a cell or organism having said mRNA to produce one or more peptide or protein products. The region containing one or more ORFs is called the coding region of the mRNA molecule. In some embodiments, the mRNA molecule further includes one or more untranslated regions (UTRs).

[0155] In some embodiments, the mRNA is a monocistronic mRNA containing only one ORF. In some embodiments, the monocistronic mRNA encodes a peptide or protein containing at least one epitope of a selected antigen (e.g., a pathogenic antigen or a tumor-associated antigen). In other embodiments, the mRNA is a polycistronic mRNA containing two or more ORFs. In some embodiments, the polycistronic mRNA encodes two or more peptides or proteins that may be the same as or different from each other. In some embodiments, each peptide or protein encoded by the polycistronic mRNA contains at least one epitope of a selected antigen. In some embodiments, the different peptides or proteins encoded by the polycistronic mRNA each contain at least one epitope of a different antigen. In any of the embodiments described in this application, the at least one epitope may be at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten epitopes of an antigen.

[0156] The term "nucleobase" encompasses purines and pyrimidines, including the natural compounds adenine, thymine, guanine, cytosine, uracil, inosine, and their natural or synthetic analogues or derivatives.

[0157] As used herein, the term "functional nucleotide analog" refers to a modified form of a classical nucleotide A, G, C, U, or T, wherein the modified form (a) retains the base-pairing properties of the corresponding classical nucleotide and (b) contains at least one chemical modification of (i) a nucleobase, (ii) a glycosyl group, (iii) a phosphate group, or (iv) any combination of (i) to (iii) of the corresponding native nucleotide. As used herein, base pairing encompasses not only classical Watson-Crick adenine-thymine, adenine-uracil, or guanine-cytosine base pairs, but also base pairs formed between a classical nucleotide and a functional nucleotide analog or between a pair of functional nucleotide analogs, wherein the arrangement of the hydrogen bond donor and hydrogen bond acceptor allows for the formation of hydrogen bonds between the modified nucleobase and a classical nucleobase or between two complementary modified nucleobase structures. For example, a functional analog of guanosine (G) retains the ability to pair with functional analogs of cytosine (C) or cytosine. An example of this type of non-classical base pairing is the base pairing between the modified nucleotide inosine and adenine, cytosine, or uracil. As described in this application, functional nucleotide analogs may be naturally occurring or non-naturally occurring. Therefore, nucleic acid molecules containing functional nucleotide analogs may have at least one modified nucleobase, glycosyl, and / or nucleoside linking. This application provides exemplary chemical modifications to the nucleobase, glycosyl, or nucleoside linking of nucleic acid molecules.

[0158] As used in this application, the term "peptide" refers to a polymer containing two to fifty (2-50) amino acid residues linked by one or more covalent peptide bonds. The term applies to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues are non-naturally occurring amino acids (e.g., amino acid analogs or non-natural amino acids).

[0159] The terms “peptide” and “protein” are used interchangeably in this application to refer to a polymer having more than fifty (50) amino acid residues linked by covalent peptide bonds. That is, the description of a peptide is equally applicable to the description of a protein, and vice versa. The term applies to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues are non-naturally occurring amino acids (e.g., amino acid analogs). As used in this application, the term covers amino acid chains of any length, including full-length proteins (e.g., antigens).

[0160] The term "antigen" refers to a substance that can be recognized by a subject's immune system (including the adaptive immune system) and can trigger an immune response (including an antigen-specific immune response) upon the subject's exposure to the antigen. In some implementations, the antigen is a protein associated with diseased cells, such as cells infected with pathogens, or neoplastic cells (e.g., tumor-associated antigens (TAAs)).

[0161] An epitope is a site on the surface of an antigen molecule that binds to a single antibody molecule, such as a localized region on the surface of an antigen capable of binding to one or more antigen-binding regions of an antibody, and which, in an animal, such as a mammal (e.g., a human), possesses antigenic or immunogenic activity and can elicit an immune response. An immunogenic epitope is the portion of a polypeptide that elicits an antibody response in an animal. An antigenic epitope is the portion of a polypeptide that binds to an antibody by any method known in this art, including, for example, by immunoassay. An antigenic epitope is not necessarily immunogenic. Epitopes typically consist of chemically active surface groups of a molecule, such as amino acids or sugar side chains, and have specific three-dimensional structural features and specific charge features. Antibody epitopes can be linear epitopes or conformational epitopes. Linear epitopes are formed from a continuous amino acid sequence in a protein. Conformational epitopes are formed from amino acids that are discontinuous in the protein sequence but are bound together when the protein folds into its three-dimensional structure. Inducible epitopes are formed when the three-dimensional structure of a protein undergoes a conformational change, such as after activation or binding of another protein or ligand. In some embodiments, an epitope is a three-dimensional surface feature of a polypeptide. In other embodiments, epitopes are linear features of the polypeptide. Generally, antigens have several or many different epitopes and can react with many different antibodies.

[0162] This application provides for the use of compounds of formula (I) or pharmaceutically acceptable salts, prodrugs, or stereoisomers thereof in targeting splenic immune cells.

[0163] G1 and G2 are each independently C2-C24 straight-chain alkylene groups that are optionally substituted with substituents;

[0164] L1 and L2 are each independently -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0165] G3 and G4 are each independently C2-C24 straight-chain alkylene groups, optionally substituted with substituents;

[0166] L3 and L4 are each independently a bond, -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-;

[0167] G5 and G6 are each independently a C2-C24 straight-chain alkyl group that is optionally substituted with a substituent or a C2-C24 straight-chain alkenyl group that is optionally substituted with a substituent.

[0168] Z is either a C1-C12 alkylene group optionally substituted with a substituent or a -R group optionally substituted with a substituent. e G7R f -;

[0169] Among them, R e and R f It is a C1-C12 alkylene group that is optionally substituted with a substituent;

[0170] G7 is -NR g -、-(3-7 membered saturated cycloalkane)-、-(3-7 membered heterocyclic alkane)-、-(3-7 membered cyclic arylene)- or -(3-7 membered cyclic heteroarylene)-;

[0171] R g C1-C12 alkylene groups optionally substituted with substituents;

[0172] X and Y are each independently a C1-C12 straight-chain alkyl group that is optionally substituted with a substituent, or -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring; R k R m and R n The C1-C12 alkyl group may be substituted by a substituent; the substituent may be selected from one or more of deuterium, tritium, halogen atom, amino, hydroxyl, cyano, nitro or alkylene.

[0173] Furthermore, when X and Z form a ring together with the nitrogen they are attached to, Z is a C3-C12 alkylene group with substituents, and the branches of X and Z form a ring together with the nitrogen they are attached to. For example, Z can be -CH2-CH2-C(CH2-CH2)-.

[0174] This application further provides the use of compounds of formula (I) above, or pharmaceutically acceptable salts, prodrugs or stereoisomers thereof, in the preparation of medicaments targeting spleen immune cells.

[0175] In one specific embodiment of the above-described uses, the immune cells are selected from lymphocytes, dendritic cells, macrophages, granulocytes, and mast cells.

[0176] This application further provides the use of compounds of formula (I) above, or pharmaceutically acceptable salts, prodrugs or stereoisomers thereof, in promoting cell polarization.

[0177] This application further provides the use of the compound of formula (I) above, or a pharmaceutically acceptable salt, prodrug or stereoisomer thereof, in the preparation of a medicament that promotes immune cell polarization.

[0178] In one specific embodiment of the use in promoting cell polarization or in the preparation of a drug that promotes the polarization of immune cells, the cell polarization is a polarization from M0 type to M1 type.

[0179] In one specific embodiment of the use in promoting cell polarization or in the preparation of a drug that promotes the polarization of immune cells, the cell polarization is a polarization from M2 type to M0 type.

[0180] In one specific embodiment of the use in promoting cell polarization or in the preparation of drugs that promote immune cell polarization, the immune cells are selected from lymphocytes, dendritic cells, macrophages, granulocytes, and mast cells. Macrophages are preferred.

[0181] In any of the uses described above, the structure of the compound of formula (I) is as follows:

[0182] In some embodiments, G1 and G2 are unsubstituted C2-C4 alkylene groups, and L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently independently substituted C2-C24 straight-chain alkyl or optionally substituted C2-C24 straight-chain alkenyl groups; Z is optionally substituted C1-C12 alkylene or optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R g Optionally substituted C1-C12 alkylene groups; wherein X and Y are each independently and optionally substituted C1-C12 straight-chain alkyl groups, -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0183] In some embodiments, L3 and L4 are bonds; G1 and G2 are each independently and optionally substituted C2-C24 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkylene groups or optionally substituted C2-C24 straight-chain alkenyl groups; Z is optionally substituted C1-C12 alkylene groups or optionally substituted -R groups. e G7R f -; where R e and R fG7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g - (3-7 membered saturated cycloalkane), - (3-7 membered heterocyclic alkane)-, - (3-7 membered cyclic arylene)- or - (3-7 membered cyclic heteroarylene)-; R g X and Y are optionally substituted C1-C12 alkylene groups; each of X and Y is independently an optionally substituted C1-C12 straight-chain alkyl group, -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0184] In some embodiments, L3 and L4 are -OC(=O)O-, and G1 and G2 are each independently and optionally substituted C2-C24 straight-chain alkylene groups; L1 and L2 are each independently and optionally -C(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkylene groups or optionally substituted C2-C24 straight-chain alkenyl groups; Z is optionally substituted C1-C12 alkylene groups or optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g - (3-7 membered saturated cycloalkane), - (3-7 membered heterocyclic alkane)-, - (3-7 membered cyclic arylene)- or - (3-7 membered cyclic heteroarylene)-; R g X and Y are optionally substituted C1-C12 alkylene groups; each of X and Y is independently an optionally substituted C1-C12 straight-chain alkyl group, -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0185] In some embodiments, G5 and G6 are optionally substituted C4-C16 straight-chain alkenyl groups, and G1 and G2 are each independently optionally substituted C2-C24 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; Z is optionally substituted C1-C12 alkylene groups, optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g - (3-7 membered saturated cycloalkane), - (3-7 membered heterocyclic alkane)-, - (3-7 membered cyclic arylene)- or - (3-7 membered cyclic heteroarylene)-; R g X and Y are optionally substituted C1-C12 alkylene groups; each of X and Y is independently an optionally substituted C1-C12 straight-chain alkyl group, -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0186] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group; G1 and G2 are each independently a optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bonded, optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bonded or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C1-C12 alkylene group or an optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R g It is an optionally substituted C1-C12 alkylene group.

[0187] In some embodiments, X and Y are each independently -G1L1G3L3G5; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bonded, optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bonded or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkylene group or an optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C1-C12 alkylene group or an optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R g It is an optionally substituted C1-C12 alkylene group.

[0188] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group, L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond, optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bond or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group, optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C1-C12 alkylene group, optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R g It is an optionally substituted C1-C12 alkylene group.

[0189] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bonded, optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bonded or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C1-C12 alkylene group or an optionally substituted -R e G7R f -; where R e and R f G7 is an optionally substituted C1-C12 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R g It is an optionally substituted C1-C12 alkylene group.

[0190] In some embodiments, Z is a C2-C4 alkylene, G1 and G2 are each independently and optionally substituted C2-C24 straight-chain alkylene, L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkyl, optionally substituted C2-C24 straight-chain alkenyl; X and Y are each independently and optionally substituted C1-C12 straight-chain alkyl, -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0191] In some implementations, Z is optionally replaced by -R e G7R f -, R e and R f G7 is an optionally substituted C1-C6 alkylene group; G7 is -NR g -or-(3-7 membered heterocyclic alkenes)-; R gThe C1-C6 alkylene groups are optionally substituted; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene groups or optionally substituted C2-C24 straight-chain alkenyl groups; X and Y are each independently optional substituted C1-C12 straight-chain alkylene groups or -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring, R k R m and R n C1-C12 alkyl groups may be substituted with substituents.

[0192] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently a optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bonded, optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bonded or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group.

[0193] In some embodiments, X and Y are each independently -G1L1G3L3G5, and Z is a C2-C4 alkylene; G1 and G2 are each independently independently - optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently - or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently - or -OC(=O)O-; G5 and G6 are each independently - optionally substituted C2-C24 straight-chain alkyl or - optionally substituted C2-C24 straight-chain alkenyl.

[0194] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are -C(=O)O-; G3 and G4 are each independently a bonded or optionally substituted C2-C24 straight-chain alkylene group; L3 and L4 are each independently a bonded or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group.

[0195] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; L3 and L4 are each independently a bond or -OC(=O)O-; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group.

[0196] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optional substituted C2-C24 straight-chain alkenyl.

[0197] In some implementations, X and Y are each independently -G1L1G3L3G5, and Z is optionally replaced by -R. e G7R f -, R e and R fG7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently substituted C2-C24 straight-chain alkylene or optionally substituted C2-C24 straight-chain alkenyl.

[0198] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is an optionally substituted -R e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optionally substituted C2-C24 straight-chain alkenyl.

[0199] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optional substituted C2-C24 straight-chain alkenyl.

[0200] In some embodiments, X and Y are each independently a C1-C3 straight-chain alkyl group, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L 2,The G3 and G4 are each independently a C2-C24 straight-chain alkylene group or optionally substituted C2-C24 straight-chain alkylene group; the L3 and L4 are each independently a C2-C24 straight-chain alkylene group or optionally substituted C2-C24 straight-chain alkenyl group; the G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkylene group or optionally substituted C2-C24 straight-chain alkenyl group.

[0201] In some embodiments, X and Y are each independently a C1-C3 straight-chain alkyl group, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L 2, The G3 and G4 are bonds; the L3 and L4 are each independently bonds or -OC(=O)O-; the G5 and G6 are each independently optional substituted C2-C24 straight-chain alkyl or optional substituted C2-C24 straight-chain alkenyl.

[0202] In some embodiments, X and Y are each independently a C1-C3 straight-chain alkyl group, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L 2, The G3 and G4 are optionally substituted C2-C24 straight-chain alkylene groups; the L3 and L4 are each independently a bond or -OC(=O)O-; the G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group.

[0203] In some embodiments, X and Y are each independently a C1-C3 straight-chain alkyl group, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L 2, The G3 and G4 are each independently a C2-C24 straight-chain alkylene group or optionally substituted C2-C24 straight-chain alkylene group; the L3 and L4 are independent C2-C24 straight-chain alkylene groups or optionally substituted C2-C24 straight-chain alkenyl groups.

[0204] In some embodiments, X and Y are each independently a C1-C3 straight-chain alkyl group, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L 2,The G3 and G4 are each independently a C2-C24 straight-chain alkylene group, which is either bonded or optionally substituted; the L3 and L4 are -OC(=O)O-; the G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkyl group or an optionally substituted C2-C24 straight-chain alkenyl group.

[0205] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently bonds or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optional substituted C2-C24 straight-chain alkenylene.

[0206] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optional substituted C2-C24 straight-chain alkenyl.

[0207] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R fG7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are bonded; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkylene or optional substituted C2-C24 straight-chain alkenyl.

[0208] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C2-C24 straight-chain alkyl or optional substituted C2-C24 straight-chain alkenyl.

[0209] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently a optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0210] In some embodiments, X and Y are each independently -G1L1G3L3G5, and Z is a C2-C4 alkylene; G1 and G2 are each independently independently -C2-C24 straight-chain alkylene with optional substitution; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently -C2-C24 straight-chain alkylene with optional substitution; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently independently -C4-C16 straight-chain alkenyl with optional substitution.

[0211] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0212] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently independently C4-C16 straight-chain alkenyl groups.

[0213] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 linear alkylene; G1 and G2 are each independently optional substituted C2-C24 linear alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded and optionally substituted C2-C24 linear alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 linear alkenyl.

[0214] In some implementations, X and Y are each independently -G1L1G3L3G5, and Z is optionally replaced by -R. e G7R f -, R e and R fG7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 linear alkylene; G1 and G2 are each independently optional substituted C2-C24 linear alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded and optionally substituted C2-C24 linear alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 linear alkenyl.

[0215] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is an optionally substituted -R e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 linear alkylene; G1 and G2 are each independently optional substituted C2-C24 linear alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded and optionally substituted C2-C24 linear alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 linear alkenyl.

[0216] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 linear alkylene; G1 and G2 are each independently optional substituted C2-C24 linear alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded and optionally substituted C2-C24 linear alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 linear alkenyl.

[0217] In some embodiments, X and Y are each independently C1-C3 straight-chain alkyl, Z is C2-C4 alkylene, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded, optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently independently optional substituted C4-C16 straight-chain alkenyl.

[0218] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently a optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0219] In some embodiments, X and Y are each independently -G1L1G3L3G5, and Z is a C2-C4 alkylene group; G1 and G2 are each independently independently -C2-C24 straight-chain alkylene groups with optional substituted structures; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently -O(=O)O-; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently independently -C4-C16 straight-chain alkenyl groups with optional substituted structures.

[0220] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0221] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0222] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 straight-chain alkenyl.

[0223] In some implementations, X and Y are each independently -G1L1G3L3G5, and Z is optionally replaced by -R. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 straight-chain alkenyl.

[0224] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is an optionally substituted -R e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 straight-chain alkenyl.

[0225] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R fG7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently optional substituted C4-C16 straight-chain alkenyl.

[0226] In some embodiments, X and Y are each independently C1-C3 straight-chain alkyl groups, Z is a C2-C4 alkylene group, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently optionally substituted C2-C24 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are bonds; L3 and L4 are each independently -OC(=O)O-; G5 and G6 are each independently independently optional substituted C4-C16 straight-chain alkenyl groups.

[0227] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently a optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently a bond; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0228] In some embodiments, X and Y are each independently -G1L1G3L3G5, and Z is a C2-C4 alkylene; G1 and G2 are each independently independently substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently a bond; and G5 and G6 are each independently independently substituted C4-C16 straight-chain alkenyl.

[0229] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently a bond; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0230] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is a C2-C4 alkylene group; G1 and G2 are each independently an optionally substituted C2-C24 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene groups; L3 and L4 are each independently a bond; and G5 and G6 are each independently an optionally substituted C4-C16 straight-chain alkenyl group.

[0231] In some embodiments, X and Y are each independently a optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded; G5 and G6 are each independently independently -substituted C4-C16 straight-chain alkenyl.

[0232] In some implementations, X and Y are each independently -G1L1G3L3G5, and Z is optionally replaced by -R. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded; G5 and G6 are each independently independently -substituted C4-C16 straight-chain alkenyl.

[0233] In some embodiments, X is an optionally substituted C1-C3 straight-chain alkyl group, Y is -G1L1G3L3G5, and Z is an optionally substituted -R e G7R f -, R e and R fG7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded; G5 and G6 are each independently independently -substituted C4-C16 straight-chain alkenyl.

[0234] In some embodiments, X is -G1L1G3L3G5, Y is an optionally substituted C1-C3 straight-chain alkyl group, and Z is an optionally substituted -R group. e G7R f -, R e and R f G7 is a -(3-7-membered heterocyclic alkylene)-, which is optionally substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are optionally substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded; G5 and G6 are each independently independently -substituted C4-C16 straight-chain alkenyl.

[0235] In some embodiments, X and Y are each independently C1-C3 straight-chain alkyl, Z is C2-C4 alkylene, and X and Z are cyclic with the nitrogen to which they are attached; G1 and G2 are each independently optional substituted C2-C24 straight-chain alkylene; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently optional substituted C2-C24 straight-chain alkylene; L3 and L4 are each independently bonded; G5 and G6 are each independently independently optional substituted C4-C16 straight-chain alkenyl.

[0236] In this application, the compound has the structure shown in formula (IA):

[0237] Among them, G1, G2, G3, G4, G5, G6, L1, L2, L3, L4, X and Y are as defined above;

[0238] G8 is an optionally substituted C1-C12 alkylene group.

[0239] In this application, the compound has the structure shown in formula (IB):

[0240] Among them, G1, G2, G3, G4, G5, G6, G7, L1, L2, L3, L4, R e R f X and Y are as defined above.

[0241] In one embodiment of the structure shown in a specific formula (IB), G1 and G2 are unsubstituted C2-C4 alkylene groups.

[0242] In one embodiment of the structure shown in the specific formula (IB), L1 and L2 are -C(=O)O-.

[0243] In one embodiment of the structure shown in Formula (IB), G3 and G4 are bonds.

[0244] In one embodiment of the structure shown in a specific formula (IB), G3 and G4 are unsubstituted C2-C4 alkylene groups.

[0245] In one embodiment of the structure shown in Formula (IB), L3 and L4 are bonds.

[0246] In one embodiment of the structure shown in the specific formula (IB), L3 and L4 are -OC (=O)O-.

[0247] In one embodiment of the structure shown in a specific formula (IB), G5 and G6 are optionally substituted C4-C16 straight-chain alkenyl groups.

[0248] In this application, the compound has the structure shown in formula (IA-1):

[0249] Where m is 1 or 3, n is 0 or 1, G5 and G6 are defined as described above, G8 is defined as described above, and G9 and G... 10 It is a C1-C5 straight-chain alkane that is optionally substituted.

[0250] In one specific embodiment, the compound has the structure shown in formula (IA-2):

[0251] Among them, G1, G2, G3, G4, G5, G6, G7, G8, L1, L2, L3, and L4 are as defined above.

[0252] In one specific embodiment, the compound has the structure shown in formula (IA-3):

[0253] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0254] In one specific embodiment, the compound has the structure shown in formula (IA-4):

[0255] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0256] In one specific embodiment, the compound has the structure shown in formula (IA-5):

[0257] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0258] In one specific embodiment, the compound has the structure shown in formula (IA-6):

[0259] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0260] In one specific embodiment, the compound has the structure shown in formula (IB-1):

[0261] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0262] In one specific embodiment, the compound has the structure shown in formula (IB-2):

[0263] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0264] In one specific embodiment, the compound has the structure shown in formula (IB-3):

[0265] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0266] In one specific embodiment, the compound has the structure shown in formula (IB-4):

[0267] Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as described above.

[0268] In one embodiment of the structure shown in formula (IA-3), G8 is an optionally substituted C2-C4 alkylene group.

[0269] In one specific embodiment of the structure shown in formula (IA-3), wherein G9 or G 10 Or both have one of the following structures: methyl, ethyl, 2-hydroxyethyl.

[0270] In one specific embodiment of the structure shown in formula (IA) and formula (IB), G5 or G6 or both have one of the following structures:

[0271] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2-chain linear alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 linear alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 linear alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7 membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C12 straight-chain alkylene, -G1L1G3L3G5 or X, Z cyclic with the nitrogen to which they are attached.

[0272] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2-chain linear alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 linear alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 linear alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7-membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C4 straight-chain alkylene or -G1L1G3L3G5.

[0273] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0274] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0275] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0276] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently hydroxyl- or amino-substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0277] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X is a C1-C4 straight-chain alkyl group; and Y is a hydroxyl-substituted C1-C4 straight-chain alkyl group.

[0278] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C2-C24 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X is G1L1G3L3G5; and Y is an amino-substituted C1-C4 straight-chain alkyl group.

[0279] In one specific embodiment, G1 and G2 are each independently a optionally substituted C2 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently a C1-C4 straight-chain alkyl group.

[0280] In one specific embodiment, G1 and G2 are each independently a optionally substituted C2 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently an optionally substituted C2-C24 straight-chain alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently G1L1G3L3G5.

[0281] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7 membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C12 straight-chain alkylene, -G1L1G3L3G5 or X, Z cyclic with the nitrogen to which they are attached.

[0282] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7-membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C4 straight-chain alkylene or -G1L1G3L3G5.

[0283] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0284] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0285] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0286] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently hydroxyl- or amino-substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0287] In one specific embodiment, G1 and G2 are each independently a optionally substituted C2 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently an optionally substituted C12-C16 straight-chain alkenyl group; Z is an optionally substituted C2-C4 alkylene group; X is a C1-C4 straight-chain alkyl group; and Y is a hydroxyl-substituted C1-C4 straight-chain alkyl group.

[0288] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently and optionally substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X is G1L1G3L3G5; and Y is an amino-substituted C1-C4 straight-chain alkyl group.

[0289] In one specific embodiment, G1 and G2 are each independently a optionally substituted C2 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently an optionally substituted C12-C16 straight-chain alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently a C1-C4 straight-chain alkyl group.

[0290] In one specific embodiment, G1 and G2 are each independently a optionally substituted C2 straight-chain alkylene group; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently an optionally substituted C12-C16 straight-chain alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently G1L1G3L3G5.

[0291] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7 membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C12 straight-chain alkylene, -G1L1G3L3G5 or X, Z cyclic with the nitrogen to which they are attached.

[0292] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently and optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently and optionally -OC(=O)O-; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group or optionally substituted -R e G7R f -; where R e and R f G1 is an optionally substituted C1-C4 alkylene; G7 is -(3-7-membered heterocyclic alkylene)-; X and Y are each independently an optionally substituted C1-C4 straight-chain alkylene or -G1L1G3L3G5.

[0293] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded or optionally substituted C2-C4 straight-chain alkylene groups; L3 and L4 are each independently bonded or -OC(=O)O-; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently independently substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0294] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0295] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently and optionally substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0296] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X and Y are each independently hydroxyl- or amino-substituted C1-C4 straight-chain alkyl groups or -G1L1G3L3G5, provided that X and Y are not simultaneously C1-C4 straight-chain alkyl groups or -G1L1G3L3G5.

[0297] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X is a C1-C4 straight-chain alkyl group; and Y is a hydroxyl-substituted C1-C4 straight-chain alkyl group.

[0298] In one specific embodiment, G1 and G2 are each independently and optionally substituted C2 straight-chain alkylene groups; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently bonded; L3 and L4 are each independently bonded; G5 and G6 are each independently C2-C4 alkyl-substituted C12-C16 straight-chain alkenyl groups; Z is an optionally substituted C2-C4 alkylene group; X is G1L1G3L3G5; and Y is an amino-substituted C1-C4 straight-chain alkyl group.

[0299] In one specific embodiment, G1 and G2 are each independently a C2 linear alkylene group that is optionally substituted; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently a C2-C4 alkyl-substituted C12-C16 linear alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently a C1-C4 linear alkyl group.

[0300] In one specific embodiment, G1 and G2 are each independently a C2 linear alkylene group that is optionally substituted; L1 and L2 are each independently -C(=O)O-; G3 and G4 are each independently a bond; L3 and L4 are each independently a bond; G5 and G6 are each independently a C2-C4 alkyl-substituted C12-C16 linear alkenyl group; Z is an optionally substituted C2-C4 alkylene group; and X and Y are each independently G1L1G3L3G5.

[0301] This application further provides the following compounds listed in Table 1, or their available salts, prodrugs, or stereoisomers:

[0302] Table 1 Representative Compounds

[0303] In any of the preceding uses, the compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof is prepared as a composition for use.

[0304] In one specific embodiment, the composition further comprises an active ingredient (which may be a therapeutic or preventative agent) and a carrier for delivering the therapeutic or preventative agent, the carrier being a cationic lipid comprising one or more compounds of formula (I) or pharmaceutically available salts thereof.

[0305] In the compositions of this application, the therapeutic or preventive agent may be a nucleic acid molecule, a small molecule compound, a polypeptide or a protein, or a mixture of two or more of them.

[0306] In one specific embodiment, the nucleic acid molecule is selected from single-stranded DNA, double-stranded DNA, short isomers, agomir, antagomir, antisense molecules, circular mRNA, small interfering RNA (siRNA), asymmetric interfering RNA (aiRNA), microRNA (miRNA), Dicersubstrate RNA (dsRNA), small hairpin RNA (shRNA), transfer RNA (tRNA), messenger RNA (mRNA), and other forms of RNA molecules known in the art, or nucleic acid mimics such as locked nucleic acid (LNA), peptide nucleic acid (PNA), and morpholine ring oligonucleotides.

[0307] In one specific embodiment, the therapeutic or preventative agent comprises at least one mRNA encoding an antigen or a fragment thereof or an epitope, preferably, the mRNA being a monocistronic mRNA or a polycistronic mRNA.

[0308] In one specific embodiment, the antigen is a tumor-specific antigen or a tumor-associated antigen.

[0309] In one specific embodiment, the mRNA comprises one or more functional nucleotide analogs, preferably selected from one or more of pseudouridine, 1-methyl-pseudouridine, or 5-methylcytosine.

[0310] In one specific embodiment, the small molecule compound is selected from one or more of the following: antitumor drugs, anti-infective drugs, local anesthetics, antidepressants, anticonvulsants, antibiotics / antibacterial agents, antifungal drugs, antiparasitic drugs, hormones, hormone antagonists, immunomodulators, neurotransmitter antagonists, antiglaucoma agents, anesthetics, or imaging agents.

[0311] As described above, the amount of carrier to therapeutic or preventative agent in the composition provided in this application is not limited. In one specific embodiment, the mass ratio of the carrier to the therapeutic or preventative agent is 1:10-50:1, i.e. (1-500):10, preferably 1.5:1. For example, it can be 1:10, 10:10, 15:10, 20:10, 30:10, 40:10, 50:10, 60:10, 70:10, 80:10, 90:10, 100:10, 150:10, 200:10, 250:10, 300:10, 350:10, 400:10, 450:10, 500:10, etc.

[0312] In some embodiments, the mass ratio of the carrier to the therapeutic or preventative agent is (10-200):10.

[0313] In some embodiments, the mass ratio of the carrier to the therapeutic or preventative agent is (10-100):10.

[0314] In some embodiments, the mass ratio of the carrier to the therapeutic or preventative agent is (10-50):10.

[0315] In one specific embodiment, the composition of this application is a nanoparticle formulation, wherein the average size of the nanoparticle formulation is 10-500 nm.

[0316] In one specific embodiment, the pKa of the nanoparticle formulation of this application is 4.5 to 8.5.

[0317] The composition provided in this application as described above, wherein the carrier is a single component comprising only cationic lipids.

[0318] The composition provided in this application as described above, wherein the carrier further comprises one or more neutral lipids, preferably selected from one or more of phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, ceramide, sterols and their derivatives.

[0319] The composition provided in this application as described above, wherein the carrier is a two-component system comprising a cationic lipid and a neutral lipid / steroid.

[0320] In one specific embodiment, the molar ratio of the cationic lipid to the neutral lipid is about 100:1 to 5:1.

[0321] In one specific embodiment, the steroid is selected from one or more of cholesterol, nonsteroidal, sitosterol, ergosterol, campesterol, stigmasterol, brassosterol, tomatine, ursolic acid, α-tocopherol, and corticosteroids.

[0322] In one specific embodiment, the molar ratio of the cationic lipid to the steroid is between 2:1 and 4:1.

[0323] As described above, the composition provided in this application further comprises one or more lipids capable of binding to a polymer. Preferably, the lipids capable of binding to the polymer are one or more selected from PEG-modified phosphatidylethanolamine, PEG-modified phosphatidic acid, PEG-modified ceramide, PEG-modified dialkylamine, PEG-modified diacylglycerol, and PEG-modified dialkylglycerol.

[0324] In one specific embodiment, the molar ratio of the cationic lipid to the lipid capable of binding to the polymer is about 100:1 to 20:1.

[0325] In one specific embodiment, the carrier in the composition of this application is a four-component system, namely cationic lipid, neutral lipid, structural lipid and polymer conjugated lipid, wherein the molar ratio of the cationic lipid, the neutral lipid, the steroid lipid and the polymer conjugated lipid is (15-70):(1-15):(15-55):(0-3).

[0326] In one specific embodiment, the composition further includes a pharmaceutically acceptable excipient.

[0327] In one specific embodiment, the excipient includes a pharmaceutically acceptable diluent.

[0328] In this application, the drug targeting spleen immune cells is a vaccine or drug used to treat or prevent tumors, viral infections, or inflammation.

[0329] The tumors can be hemangiomas, lymphangiomas, hamartomas, fibromas, lipomas, primary malignant tumors of the spleen, metastatic tumors of the spleen, etc.

[0330] The viral infections mentioned can be hepatitis viruses, cytomegalovirus (CMV), Epstein-Barr virus (EBV), human immunodeficiency virus (HIV), parvovirus B19, influenza virus infection, measles virus, rubella virus, epidemic hemorrhagic fever virus, infectious spleen and kidney necrosis virus (ISKNV), etc.

[0331] Inflammation can manifest as acute inflammatory splenomegaly, necrotizing splenomegaly, chronic splenomegaly, splenic abscess, splenic tuberculosis, immune splenomegaly, hematologic disorders of splenomegaly, splenic infarction, etc.

[0332] Example 1: Preparation of the compound

[0333] Example 1-1 Synthesis of Compound 9

[0334] Synthetic route

[0335] Step 1: Synthesis of intermediate 9-2

[0336] To a mixture of geraniol (5 g, 16.86 mmol, 1 eq) and triethylamine (2.33 mL, 16.86 mmol, 1 eq) in dichloromethane (100 mL), p-nitrophenyl chloroformate (4.07 g, 20.03 mmol, 1.2 eq) dissolved in dichloromethane was slowly added dropwise. The reaction mixture was stirred at room temperature for 3 h, and the reaction was stopped. The mixture was extracted with water, and the combined organic layers were dried over MgSO4 and the solvent was removed under vacuum to give crude product 9-1. Hydroxybutyl acrylate (2.11 g, 20.03 mmol, 1.2 eq), potassium carbonate (2.76 g, 20.03 mmol, 1.2 eq), and N,N-dimethylformamide solvent (200 mL) were added to a 250 mL round-bottom flask containing crude product 9-1. The mixture was stirred at 80 °C for 3 h, and TLC showed that compound 9-1 completely disappeared. After removing DMF under vacuum, the product was washed with brine, and the combined organic layers were dried with MgSO4 and the solvent was removed under vacuum to obtain a crude product. The crude product was purified by column chromatography (elution buffer: PE / EA = 10 / 1), and the pure product fraction was evaporated to obtain a pale yellow oily compound 9-2 (10.06 g, 69%).

[0337] Step 2: Synthesis of Compound 9

[0338] N,N-dimethylethylenediamine (30 μL, 0.33 mmol, 1 eq) and compound 9-2 (300 μL, 1.01 mmol, 3 eq) were mixed and stirred at 70 °C for 48 h. The mixture was purified by column chromatography (silica gel column, eluent was dichloromethane solution containing 0-10% methanol (v / v)) to give compound 9 (370 mg, 54%) as a pale yellow oil. 1HNMR (400MHz, CDCl3) δ5.38(t,J=6.9Hz,2H),5.08(t,J=6.6Hz,2H),4.65(d,J=7.2Hz,4H),4.13(dt,J=26.9,5.9Hz,8H),2.79(t,J=7.1Hz, 4H),2.62(d,J=6.6Hz,2H),2.45(t,J=7.1Hz,6H),2.34(s,6H),2.12-2.02(m,8H),1.80-1.72(m,8H),1.70(d,J=15.4Hz,12H),1.60(s,6H).

[0339] Synthesis of Compound 32 in Examples 1-2

[0340] Compound 32 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.38 (dd, J = 7.2, 6.1 Hz, 2H), 5.10 (dd, J = 7.2, 2.5 Hz, 4H), 4.67 (d, J = 7.2 Hz, 4H), 4.36–4.27 (m, 8H), 2.89–2.60 (m, 4H), 2.49–2.40 (m, 8H), 2.32–1.79 (m, 24H), 1.73–1.67 (m, 12H), 1.61–1.47 (m, 14H).

[0341] Synthesis of Compound 21 in Examples 1-3

[0342] Compound 21 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.08 (td, J = 7.1, 1.2 Hz, 3H), 4.34–4.29 (m, 12H), 4.21–4.16 (m, 6H), 2.74 (d, J = 7.0 Hz, 6H), 2.49–2.25 (m, 24H), 1.98 (dt, J = 21.2, 7.3 Hz, 6H), 1.75–1.67 (m, 14H), 1.60–1.54 (m, 13H), 1.41–1.11 (m, 6H), 0.92 (d, J = 6.1 Hz, 9H).

[0343] Synthesis of Compound 8 in Examples 1-4

[0344] Compound 8 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.38 (t, J = 7.2 Hz, 2H), 5.07 (d, J = 6.8 Hz, 2H), 4.67 (d, J = 7.2 Hz, 4H), 4.31 (ddd, J = 8.2, 5.1, 2.1 Hz, 8H), 2.75 (ddd, J = 48.6, 13.4, 6.5 Hz, 4H), 2.49–2.41 (m, 8H), 2.30 (s, 2H), 2.07 (dd, J = 11.8, 6.5 Hz, 8H), 1.95 (s, 2H), 1.72–1.67 (m, 12H), 1.60 (s, 6H), 1.52 (s, 3H), 1.27 (d, J = 12.2 Hz, 3H).

[0345] Synthesis of Compound 10 in Examples 1-5

[0346] Compound 10 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.37 (dd, J = 7.8, 6.6 Hz, 2H), 5.08 (t, J = 6.7 Hz, 2H), 4.65 (d, J = 7.2 Hz, 4H), 4.12 (dt, J = 27.8, 6.0 Hz, 8H), 2.75 (t, J = 7.1 Hz, 4H), 2.42 (dq, J = 14.0, 7.3 Hz, 8H), 2.31 (s, 6H), 2.14–2.03 (m, 8H), 1.78–1.72 (m, 8H), 1.72 (s, 6H), 1.68 (s, 6H), 1.67–1.62 (m, 2H), 1.60 (s, 6H).

[0347] Synthesis of Compound 33 in Examples 1-6

[0348] The synthesis route is as follows:

[0349] 33.1 Preparation of intermediate 33-1

[0350] Farnesol (10 g, 45.25 mmol, 1 eq) was dissolved in 50 mL of dichloromethane, and acryloyl chloride (54.30 mmol, 1.2 eq) and triethylamine (67.88 mmol, 1.5 eq) were added dropwise. The reaction was carried out overnight, and the mixture was purified by column chromatography (PE / EA = 10:1) to give intermediate 33-1. ¹H NMR (400 MHz, CDCl₃) δ 6.46–6.34 (m, ¹H), 6. 12(dd,J=17.3,10.4Hz,1H),5.86-5.76(m,1H),5.40(t,J=7.0Hz,1H),5.17-5.03(m,2H),4.66 (d,J=7.3Hz,2H),2.16-2.03(m,6H),2.00-1.95(m,2H),1.78(s,3H),1.68(s,3H),1.60(s,6H).

[0351] 33.2 Preparation of Compound 33

[0352] N,N-dimethylethylenediamine (32 μL, 0.36 mmol, 1 eq) and compound 33-1 (300 μL, 1.09 mmol, 3 eq) were mixed and stirred at 70 °C for 48 h. The mixture was purified by column chromatography (silica gel column, eluent was dichloromethane solution containing 0-10% methanol (v / v)) to give compound 33 (380 mg, 60%) as a pale yellow oil. 1HNMR (400MHz, CDCl3) δ5.34(t,J=7.2Hz,2H),5.14-5.04(m,4H),4.56(d,J=7.2Hz,4H),2.79(t,J=7.2Hz,4H),2.60(t,J=7. 1Hz,2H),2.45(t,J=7.2Hz,6H),2.30(s,6H),2.13-2.04(m,12H),1.99-1.95(m,4H),1.76(s,6H),1.68(s,6H),1.60(s,12H).

[0353] Synthesis of Compound 16 in Examples 1-7

[0354] Compound 16 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.37 (t, J = 7.2 Hz, 2H), 5.08 (t, J = 6.6 Hz, 2H), 4.65 (d, J = 7.2 Hz, 4H), 4.12 (dt, J = 27.1, 5.9 Hz, 8H), 2.87–2.67 (m, 4H), 2.49–2.34 (m, 9H), 2.14–2.00 (m, 9H), 1.92 (dd, J = 19.8, 12.4 Hz, 3H), 1.79–1.69 (m, 16H), 1.68 (s, 6H), 1.60 (s, 6H), 1.46 (d, J = 17.4 Hz, 3H).

[0355] Synthesis of Compound 30 in Examples 1-8

[0356] Compound 30 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.38 (s, 2H), 5.09 (s, 4H), 4.66 (d, J = 7.0 Hz, 4H), 4.30 (d, J = 11.8 Hz, 8H), 2.72 (dd, J = 41.3, 32.6 Hz, 12H), 2.43 (d, J = 6.6 Hz, 4H), 2.19–1.97 (m, 18H), 1.70 (d, J = 17.8 Hz, 12H), 1.59 (s, 12H), 1.43–0.89 (m, 10H).

[0357] Synthesis of Compound 15 in Examples 1-9

[0358] Compound 15 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.38 (t, J = 7.1 Hz, 4H), 5.08 (t, J = 6.3 Hz, 4H), 4.65 (d, J = 7.2 Hz, 8H), 4.12 (dt, J = 28.2, 5.6 Hz, 18H), 2.75 (t, J = 7.1 Hz, 8H), 2.42 (t, J = 7.2 Hz, 16H), 2.13–1.99 (m, 18H), 1.73 (dd, J = 6.2, 3.2 Hz, 30H), 1.68 (s, 15H), 1.60 (s, 15H).

[0359] Synthesis of Compound 2 in Examples 1-10

[0360] Compound 2 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.43–5.27 (m, 2H), 5.08 (s, 2H), 4.67 (d, J = 7.2 Hz, 2H), 4.60 (dd, J = 13.8, 7.0 Hz, 2H), 4.28 (dt, J = 36.3, 19.1 Hz, 6H), 2.74 (dd, J = 21.3, 14.6 Hz, 5H), 2.62 (s, 6H), 2.44 (ddd, J = 32.9, 16.7, 9.9 Hz, 6H), 2.13–2.02 (m, 8H), 1.88 (s, 2H), 1.73–1.67 (m, 12H), 1.60 (s, 6H), 1.27 (d, J = 12.3 Hz, 3H).

[0361] Synthesis of Compound 51 in Examples 1-11

[0362] Compound 51 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.37 (t, J = 7.2 Hz, 4H), 4.67 (d, J = 7.2 Hz, 8H), 4.31 (dd, J = 13.2, 5.7 Hz, 16H), 2.79 (t, J = 6.9 Hz, 8H), 2.48 (s, 17H), 2.00 (t, J = 6.2 Hz, 8H), 1.71 (s, 12H), 1.52 (dt, J = 13.2, 6.6 Hz, 4H), 1.46–1.32 (m, 18H), 1.32–1.18 (m, 32H), 1.09 (dddd, J = 14.6, 12.1, 10.3, 7.1 Hz, 26H), 0.90–0.81 (m, 48H).

[0363] Synthesis of Compound 56 in Examples 1-12

[0364] Compound 56 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.36 (t, J = 6.8 Hz, 2H), 4.67 (d, J = 7.2 Hz, 3H), 4.34–4.27 (m, 6H), 2.83 (dd, J = 13.6, 6.9 Hz, 2H), 2.63 (dd, J = 13.8, 7.4 Hz, 6H), 2.52–2.41 (m, 6H), 2.22–1.78 (m, 9H), 1.70 (d, J = 8.4 Hz, 8H), 1.60–0.97 (m, 42H), 0.85 (dd, J = 8.5, 6.7 Hz, 24H).

[0365] Synthesis of Compound 26 in Examples 1-13

[0366] Compound 26 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.42–5.28 (m, 2H), 5.08 (s, 4H), 4.74–4.51 (m, 4H), 4.30 (dd, J = 15.2, 5.7 Hz, 6H), 2.71 (d, J = 6.5 Hz, 4H), 2.65–2.37 (m, 12H), 2.31 (s, 4H), 2.12–2.01 (m, 12H), 1.99–1.90 (m, 4H), 1.73–1.65 (m, 12H), 1.59 (s, 12H), 1.28 (m, 2H).

[0367] Synthesis of Compound 25 in Examples 1-14

[0368] Compound 25 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.38 (t, J = 7.2 Hz, 2H), 5.09 (s, 4H), 4.67 (d, J = 7.2 Hz, 4H), 4.31 (dd, J = 12.9, 5.7 Hz, 8H), 2.79 (t, J = 7.0 Hz, 4H), 2.64 (s, 2H), 2.49 (t, J = 7.0 Hz, 6H), 2.35 (s, 6H), 2.08 (dt, J = 16.2, 6.9 Hz, 12H), 1.99–1.94 (m, 4H), 1.72 (s, 6H), 1.68 (s, 6H), 1.60 (s, 12H).

[0369] Synthesis of Compound 29 in Examples 1-15

[0370] Compound 29 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.34 (t, J = 7.2 Hz, 3H), 5.15–5.03 (m, 6H), 4.56 (d, J = 7.2 Hz, 6H), 2.75 (t, J = 7.4 Hz, 6H), 2.45–2.32 (m, 14H), 2.28 (s, 6H), 2.08 (dt, J = 15.6, 5.8 Hz, 18H), 2.01–1.95 (m, 6H), 1.76 (s, 9H), 1.70–1.64 (m, 11H), 1.63–1.52 (m, 20H).

[0371] Synthesis of Compound 5 in Examples 1-16

[0372] Compound 5 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃): δ 5.37 (t, J = 7.0 Hz, 3H), 5.07 (d, J = 6.6 Hz, 3H), 4.67 (d, J = 7.2 Hz, 6H), 4.31 (dd, J = 13.5, 5.6 Hz, 12H), 2.75 (t, J = 7.1 Hz, 6H), 2.66 (s, 3H), 2.56–2.33 (m, 19H), 2.15–1.99 (m, 14H), 1.73–1.67 (m, 18H), 1.60 (s, 9H).

[0373] Synthesis of Compound 42 in Examples 1-17

[0374] Compound 42 was prepared using the method described in Example 33. ¹H NMR (400 MHz, CDCl₃) δ 5.32 (d, J = 7.1 Hz, 2H), 4.59 (d, J = 7.1 Hz, 4H), 2.76 (t, J = 7.0 Hz, 4H), 2.48–2.33 (m, 8H), 2.30 (s, 6H), 2.00 (s, 4H), 1.67 (d, J = 14.0 Hz, 8H), 1.52 (dd, J = 13.2, 6.6 Hz, 2H), 1.22 (ddd, J = 34.9, 33.1, 22.4 Hz, 36H), 0.86 (t, J = 6.4 Hz, 24H).

[0375] Synthesis of Compound 53 in Examples 1-18

[0376] Compound 53 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.36 (d, J = 7.4 Hz, 3H), 4.67 (d, J = 7.2 Hz, 3H), 4.29 (dd, J = 15.6, 6.4 Hz, 9H), 2.74 (d, J = 3.4 Hz, 6H), 2.60 (s, 6H), 2.52–2.35 (m, 12H), 2.00 (d, J = 6.0 Hz, 6H), 1.70 (d, J = 15.6, 6.4 Hz, 9H). J=9.2Hz,10H),1.51(dt,J=19.8,6.6Hz,6H),1.37(ddd,J=15.7,11.6,4.4Hz,16H),1.30-1. 20(m,26H),1.18-1.11(m,8H),1.07(ddd,J=14.9,11.5,5.9Hz,12H),0.86(t,J=7.6Hz,36H).

[0377] Synthesis of Compound 52 in Examples 1-19

[0378] Compound 52 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.37 (t, J = 7.2 Hz, 4H), 4.67 (d, J = 7.2 Hz, 8H), 4.31 (dd, J = 13.9, 5.6 Hz, 16H), 2.74 (s, 8H), 2.46 (t, J = 6.7 Hz, 12H), 2.00 (d, J = 6.4 Hz, 10H), 1.71 (s, 12H), 1 .65(s,6H),1.52(dd,J=13.2,6.7Hz,4H),1.38(dd,J=18.1,13.7Hz,18H),1.32-1.20(m ,32H),1.14(dd,J=11.4,4.5Hz,9H),1.11-1.01(m,16H),0.86(dd,J=8.3,6.8Hz,48H).

[0379] Synthesis of Compound 13 in Examples 1-20

[0380] Compound 13 was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.37 (t, J = 7.2 Hz, 3H), 5.08 (t, J = 6.5 Hz, 3H), 4.65 (d, J = 7.2 Hz, 6H), 4.17–4.07 (m, 12H), 2.75 (t, J = 7.2 Hz, 6H), 2.53–2.29 (m, 20H), 2.12–2.02 (m, 12H), 1.78–1.73 (m, 11H), 1.72 (s, 12H), 1.68 (s, 10H), 1.60 (s, 8H), 1.57–1.52 (m, 2H).

[0381] Synthesis of Compound 50 in Examples 1-21

[0382] The compound was prepared using the method described in Example 1. ¹H NMR (400 MHz, CDCl₃) δ 5.37 (t, J = 7.0 Hz, 2H), 4.67 (d, J = 7.2 Hz, 4H), 4.31 (qd, J = 6.3, 2.6 Hz, 8H), 2.74 (t, J = 6.8 Hz, 4H), 2.49–2.35 (m, 12H), 2.00 (t, J = 6.6 Hz, 6.6 Hz, 8H). Hz,6H),1.71(s,6H),1.53(dd,J=13.3,6.7Hz,2H),1.35(dd,J=14.5,6.1Hz,8H),1.31-1 .19(m,16H),1.09(dddd,J=14.8,12.4,10.5,7.1Hz,14H),0.85(dd,J=8.7,6.7Hz,24H).

[0383] Example 2: Preparation of Nanoparticle Composition

[0384] Twenty-one cationic lipid compounds prepared in Example 1 were dissolved in anhydrous ethanol at a molar ratio of 66.67:25.67:6.67:1.00 with cholesterol (Avetrol (Shanghai) Pharmaceutical Technology Co., Ltd.), DSPC alcohol (Avetrol (Shanghai) Pharmaceutical Technology Co., Ltd.), and DMG-PEG2000 (Avetrol (Shanghai) Pharmaceutical Technology Co., Ltd.). Lipid nanoparticles (LNPs) were prepared at a weight ratio of approximately 10:1 cationic lipids to firefly luciferase (luc) mRNA. Briefly, the mRNA was diluted in 25 mM sodium acetate solution (pH 5.2), and the ethanol solution of lipids was mixed with the aqueous solution of mRNA at a ratio of approximately 1:3 (volume / volume) using a syringe pump at a total flow rate of 12 mL / min. The LNPs were replaced with ddH2O by dialysis to remove the ethanol. Finally, the lipid nanoparticles were filtered through a sterile filter with 0.2 μm pores to obtain the LNP formulation encapsulating firefly luciferase mRNA (LNP-mluc).

[0385] Example 3: In vivo evaluation of luciferase mRNA using lipid nanoparticle composition

[0386] The lipid nanoparticle composition (LNP-mluc) prepared in Example 2 was evaluated in vivo for luciferase mRNA. The luc mRNA from Shanghai Hexincheng Biotechnology expresses the luciferase protein and was initially isolated from fireflies. luc is commonly used in mammalian cell cultures to measure gene expression and cell viability. It emits bioluminescence in the presence of its substrate, luciferin. The study was conducted on 5-6 week old female Balb / c mice (Shanghai Slack Laboratory Animal Co., Ltd.) according to guidelines established by the Laboratory Animal Control Committee (ACC) and the Canadian Council for Animal Control (CCAC). Six hours after administration, mice were intraperitoneally injected with 100 μL of 30 mg / mL D-luciferin potassium salt (Adamas Reagent Co., Ltd.). Imaging of the mice was performed 10 minutes later using a PerkinElmer in vivo imaging system. The compound exhibiting strong fluorescence intensity in the liver was selected. Compounds 9, 32, 21, 8, 10, 33, 16, 30, 15, 2, 51, 56, 26, 25, 29, 5, 42, 53, 52, 13, and 50 exhibit strong fluorescence signals in the spleen, with fluorescence intensities generally above approximately 1.17E+07. In particular, compound 50 has the highest fluorescence intensity, reaching 1.03E+08. Compound 17, which has a strong signal in the lungs, has a fluorescence signal intensity of approximately 6.95E+05.

[0387] Example 4: Preparation method of the four-component LNP system

[0388] Compound 50, prepared in Example 1, yielded an LNP based on a traditional four-component system that primarily targets the spleen. The preparation method is as follows:

[0389] 1. Prepare 2.5μl, 20μl, 200μl, and 1000μl pipettes; 10, 200, and 1000μl nuclease-free sterile pipette tips; 1.5ml centrifuge tubes; 15ml centrifuge tubes; DEPC water; sodium acetate buffer (pH=5.2); 75% alcohol; anhydrous ethanol; 1.5ml centrifuge tube racks; and 15 / 50ml centrifuge tube racks (all liquids and tubes mentioned above are nuclease-free and sterile).

[0390] 2. Prepare working solutions of cholesterol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), DSPC alcohol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), cationic lipids, and DMG-PEG2000 (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.) in advance with anhydrous ethanol. The concentration of each working solution is 10 mg / ml. Seal and store in the refrigerator to prevent ethanol evaporation from affecting the concentration.

[0391] 3. According to the mass ratio of cationic lipid (compound 50), cholesterol, DSPC, and DMG-PEG 2000 of 50:38.5:10:1.5, take cationic lipid, cholesterol, DSPC, and DMG-PEG and mix them in a 1.5ml centrifuge tube. Add a certain volume of anhydrous ethanol to prepare a final concentration of 5mg / ml mix. Vortex for 1min to mix, sonicate for 1min, and continue vortexing for 1min. Separately, take 3 volumes of 25mM sodium acetate solution (pH 5.2) and add the mix dropwise to the vortexed sodium acetate buffer to obtain LNP.

[0392] Example 5: Investigation of the mass ratio of LNP to RNA in a four-component system (Preparation of LNP-mRNA complex 1)

[0393] The LNP prepared in Example 4 was placed in a dialysis bag (3.5K, Thermo Scientific) and dialyzed with ultrapure water at room temperature for more than 3 hours. The concentration of LNP after dialysis was between 0.8-1 μg / μl. The LNP:RNA (w / w) was mixed at ratios of 10:1, 7.5:1, 5:1 and 3:1 and incubated for 15 min. The mixture was then diluted with physiological saline (150mM Nacl) to 200 μl to prepare complex 1 for tail vein injection, with an injection volume of 200 μl / mouse.

[0394] Example 6: Preparation and formulation optimization of the three-component cholesterol-removing system LNP

[0395] 1. Prepare 2.5μl, 20μl, 200μl, and 1000μl pipettes; 10, 200, and 1000μl nuclease-free sterile pipette tips; 1.5ml centrifuge tubes; 15ml centrifuge tubes; DEPC water; sodium acetate buffer (pH=5.2); 75% alcohol; anhydrous ethanol; 1.5ml centrifuge tube racks; and 15 / 50ml centrifuge tube racks (all liquids and tubes mentioned above are nuclease-free and sterile).

[0396] 2. Prepare working solutions of DSPC alcohol (Aivert (Shanghai) Pharmaceutical Technology Co., Ltd.), cationic lipids, and DMG-PEG2000 (Aivert (Shanghai) Pharmaceutical Technology Co., Ltd.) in advance with anhydrous ethanol. The concentration of each working solution is 10 mg / ml. Seal and store in a refrigerator to prevent ethanol evaporation from affecting the concentration.

[0397] 3. According to the three-component molar ratio shown in Table 1 below, take cationic lipid (compound 50), DSPC, and DMG-PEG and mix them in a 1.5ml centrifuge tube. Add a certain volume of anhydrous ethanol to prepare a final concentration of 5mg / ml mix. Vortex for 1min to mix, sonicate for 1min, and continue vortexing for 1min. Separately, take 3 times the volume of 25mM sodium acetate solution (pH 5.2) and add the mix dropwise into the vortexed sodium acetate buffer to obtain the three-component LNP with cholesterol removal.

[0398] Table 1 shows the particle size and zeta potential of the three-component LNP formulation in Example 6.

[0399] Example 7: Preparation of LNP-mRNA complex 2

[0400] The LNP prepared in Example 6 was placed in a dialysis bag (3.5K, Thermo Scientific) and dialyzed with ultrapure water at room temperature for more than 3 hours. The concentration of LNP after dialysis was between 0.8-1 μg / μl. The LNP:RNA (w / w) was mixed at a ratio of 10:1 and incubated for 15 min. It was then diluted with physiological saline (150mM Nacl) to 200 μl to prepare complex 2 for tail vein injection, with an injection volume of 200 μl / mouse.

[0401] Example 8: Preparation and Formulation Optimization of a Three-Component LNP System with PEG Removal

[0402] 1. Prepare 2.5μl, 20μl, 200μl, and 1000μl pipettes; 10, 200, and 1000μl nuclease-free sterile pipette tips; 1.5ml centrifuge tubes; 15ml centrifuge tubes; DEPC water; sodium acetate buffer (pH=5.2); 75% alcohol; anhydrous ethanol; 1.5ml centrifuge tube racks; and 15 / 50ml centrifuge tube racks (all liquids and tubes mentioned above are nuclease-free and sterile).

[0403] 2. Prepare working solutions of cholesterol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), DSPC alcohol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), and cationic lipids in advance using anhydrous ethanol. The concentration of each working solution is 10 mg / ml. Seal and store in a refrigerator to prevent ethanol evaporation from affecting the concentration.

[0404] 3. According to the three-component molar ratio shown in Table 2 below, take cationic lipid (compound 50), cholesterol, and DSPC and mix them in a 1.5ml centrifuge tube. Add a certain volume of anhydrous ethanol to prepare a final concentration of 5mg / ml mix. Vortex for 1min to mix, sonicate for 1min, and continue vortexing for 1min. Separately, take 3 times the volume of 25mM sodium acetate solution (pH 5.2) and add the mix dropwise into the vortexed sodium acetate buffer to obtain the three-component LNP with PEG removed.

[0405] Table 2 shows the particle size and zeta potential of the three-component LNP formulation in Example 8.

[0406] Example 9: Preparation of LNP-mRNA complex 3

[0407] The LNP prepared in Example 8 was placed in a dialysis bag (3.5K, Thermo Scientific) and dialyzed with ultrapure water at room temperature for more than 3 hours. The LNP concentration after dialysis was between 0.8-1 μg / μl. The LNP:RNA (w / w) was mixed at a ratio of 10:1 and incubated for 15 min. It was then diluted with physiological saline (150mM Nacl) to 200 μl to prepare complex 3 for tail vein injection, with an injection volume of 200 μl / mouse.

[0408] Example 10: Preparation and Formulation Optimization of the Two-Component LNP System

[0409] 1. Prepare 2.5μl, 20μl, 200μl, and 1000μl pipettes; 10, 200, and 1000μl nuclease-free sterile pipette tips; 1.5ml centrifuge tubes; 15ml centrifuge tubes; DEPC water; sodium acetate buffer (pH=5.2); 75% alcohol; anhydrous ethanol; 1.5ml centrifuge tube racks; and 15 / 50ml centrifuge tube racks (all liquids and tubes mentioned above are nuclease-free and sterile).

[0410] 2. Prepare working solutions of cationic lipids (compound 50), DOPE alcohol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), DSPC alcohol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.), and cholesterol (Avetuo (Shanghai) Pharmaceutical Technology Co., Ltd.) in advance with anhydrous ethanol. The concentration of each working solution is 10 mg / ml. Seal and store in the refrigerator to prevent ethanol evaporation from affecting the concentration.

[0411] 3. According to the mass ratio of cationic lipids (compound 50) to DOPE or cholesterol or DPSC of 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, and 1:5, mix the cationic lipids and DOPE or cholesterol or DPSC in a 1.5 ml centrifuge tube, add a certain volume of anhydrous ethanol to prepare a final concentration of 5 mg / ml mix, vortex for 1 min to mix, sonicate for 1 min, and continue vortexing for 1 min; separately, take 3 volumes of 25 mM sodium acetate solution (pH 5.2), and add the mix dropwise to the vortexed sodium acetate buffer to obtain LNP.

[0412] Example 11: Preparation of LNP-mRNA complex 4

[0413] The LNP prepared in Example 10 was placed in a dialysis bag (3.5K, Thermo Scientific) and dialyzed with ultrapure water at room temperature for more than 3 hours. The concentration of LNP after dialysis was between 0.8-1 μg / μl. The LNP:RNA (w / w) was mixed at a ratio of 10:1 and incubated for 15 minutes to obtain complex 4 for tail vein injection.

[0414] Example 12: Preparation of the single-component system LNP

[0415] 1. Prepare 2.5μl, 20μl, 200μl, and 1000μl pipettes; 10, 200, and 1000μl nuclease-free sterile pipette tips; 1.5ml centrifuge tubes; 15ml centrifuge tubes; DEPC water; sodium acetate buffer (pH=5.2); 75% alcohol; anhydrous ethanol; 1.5ml centrifuge tube racks; and 15 / 50ml centrifuge tube racks (all liquids and tubes mentioned above are nuclease-free and sterile).

[0416] 2. Prepare cationic lipid working solutions with anhydrous ethanol (using compounds 50, 53, and 56 respectively), with a concentration of 5 mg / ml. Seal and store in a refrigerator to prevent ethanol evaporation from affecting the concentration.

[0417] 3. Take 3 times the volume of 25mM sodium acetate solution (pH 5.2) and add the previously prepared cationic lipid working solution dropwise into the sodium acetate buffer solution under vortex conditions to obtain the single-component system LNP (single-component system LNP-50, single-component system LNP-53, single-component system LNP-56).

[0418] Example 13: Investigation of the mass ratio of LNP to RNA in a single-component system (Preparation of LNP-mRNA complex 5)

[0419] The single-component system LNP-50 prepared in Example 12 was placed in a dialysis bag (3.5K, Thermo Scientific) and dialyzed with ultrapure water at room temperature for more than 3 hours. The LNP concentration after dialysis was between 0.8-1 μg / μl. LNP:RNA (w / w) was mixed and incubated for 15 min at ratios of 15:1, 10:1, 7.5:1, 5:1 and 3:1, 1.5:1, 1.5:2. The mixture was then diluted to 200 μl with physiological saline (150 mMnAcl) to prepare complex 5 for tail vein injection, with an injection volume of 200 μl / mouse.

[0420] Example 14: Investigation into the relationship between the mass ratio of LNP to RNA in a four-component system and the stability of the LNP-mRNA complex.

[0421] LNP-luc-mRNA complex 1 was prepared according to the method in Example 5. Its particle size and zeta potential were detected using a Malvern Panaco nanoparticle size potentiostat to reflect its physicochemical properties and stability. The results are shown in Figures 1-1 and 1-2. The complex formed under the condition of 5:1 mass ratio is unstable and forms large aggregates immediately after preparation. Stable LNP-RNA is formed under the other ratios, with a particle size in the range of 200nm-250nm and a zeta potential that transitions from positive to negative.

[0422] Example 15: Investigation into the relationship between the mass ratio of LNP to RNA in a four-component system and targeting in wild-type Balb / C mice

[0423] LNP-luc-mRNA complex 1 was prepared according to the method in Example 5. 5 μg mRNA / mouse was injected via the tail vein. Six hours later, the biofluorescence intensity distribution in the mouse body and the biofluorescence intensity distribution in the dissected heart, liver, spleen, lungs, kidneys, and bone marrow were measured using an IVIS small animal in vivo imaging system. Before imaging, mice were intraperitoneally injected with 100 μL of D-luciferin potassium salt (30 mg / mL, dissolved in PBS). The results are shown in Figures 2-1 and 2-2. A spleen targeting ratio of 10:1 showed optimal targeting.

[0424] Example 16: Stability Study of Optimized Three-Component LNP Formulation for Cholesterol Removal

[0425] LNP-luc-mRNA complex 2 was prepared according to the method in Example 7. Its particle size and zeta potential were detected using a Malvern Panaco nanoparticle size potentiometer to reflect its physicochemical properties and stability. The results are shown in Table 1. None of the formulations met the requirements of a stable complex with a particle size of less than 200 nm and a PDI of less than 0.2.

[0426] Example 17: Stability Study of Optimized Three-Component LNP Formulation with PEG Removal

[0427] LNP-luc-mRNA complex 3 was prepared according to the method in Example 9. Its particle size and zeta potential were detected using a Malvern Panaco nanoparticle size potentiometer to reflect its physicochemical properties and stability. The results are shown in Table 2. None of the formulations met the requirements of a stable complex with a particle size of less than 200 nm and a PDI of less than 0.2.

[0428] Example 18: Stability Study of LNP Formulation Optimization in Two-Component Systems

[0429] LNP-luc-mRNA complex 4 was prepared according to the method in Example 11. Its particle size and zeta potential were detected using a Malvern Panaco nanoparticle size potentiostat to reflect its physicochemical properties and stability. The results are shown in Figures 3-1 and 3-2. The two-component systems formed with cholesterol or DSPC were unstable and formed large aggregates immediately after preparation. As shown in Figures 4-1 and 4-2, the complexes formed with DOPE at mass ratios of 5:1, 4:1, and 2:1 were unstable and formed large aggregates immediately after preparation. Stable LNP-RNA was formed at the other ratios, with a particle size in the range of 200nm-300nm and a zeta potential that transitioned from positive to negative.

[0430] Example 19: Targeting Study of Optimized Two-Component LNP Formulation in Wild-Type Balb / C Mice

[0431] LNP-luc-mRNA complex 4 was prepared according to the method in Example 11. 5 μg mRNA / mouse was injected via tail vein. Six hours later, the biofluorescence intensity distribution in the mouse body and the biofluorescence intensity distribution in the dissected heart, liver, spleen, lungs, kidneys, and bone marrow were measured using an IVIS small animal in vivo imaging system. Before imaging, the mouse was intraperitoneally injected with 100 μL of D-luciferin potassium salt (30 mg / mL, dissolved in PBS). The results are shown in Figures 5-1 and 5-2. Spleen showed strong targeting at a mass ratio of 1:(2-5).

[0432] Example 20: Investigation into the relationship between the mass ratio of LNP to RNA in a single-component system and the stability of the LNP-mRNA complex.

[0433] LNP-luc-mRNA complex 5 was prepared according to the method in Example 13. Its particle size and zeta potential were detected using a Malvern Panaco nanoparticle size potentiostat to reflect its physicochemical properties and stability. The results are shown in Figures 6-1 and 6-2. The complexes formed under mass ratios of 3:1, 1.5:1, 15:1 and 1.5:2 were stable, with particle sizes in the range of 200 nm to 300 nm and a zeta potential that transitioned from positive to negative.

[0434] Example 21: Investigation into the relationship between the mass ratio of LNP to RNA in a single-component system and targeting in wild-type Balb / C mice

[0435] The LNP-luc-mRNA complex 5 was prepared according to the method in Example 13. 5 μg mRNA / mouse was injected via tail vein. Six hours later, the biofluorescence intensity distribution in the mouse body and the biofluorescence intensity distribution in the dissected heart, liver, spleen, lungs, kidneys, and bone marrow were measured using an IVIS small animal in vivo imaging system. Mice were injected intraperitoneally with 100 μL of D-luciferin potassium salt (30 mg / mL, dissolved in PBS) before imaging. The results are shown in Figures 7 and 8. When the LNP to RNA mass ratio was 15:1, there was still high signal expression in the liver. When the LNP to RNA mass ratio was 3:1 and 1.5:1, the spleen showed better targeting (81.2%), with similar signal intensities. Finally, a formulation with an LNP to RNA mass ratio of 1.5:1 was selected for subsequent experiments. This formulation reduced the amount of LNP used in delivering nucleic acid drugs and improved safety.

[0436] Example 22: In vivo targeting validation of single-component system formulations in other spleen-targeting LNPs

[0437] LNP-luc-mRNA complexes 53 and 56 were prepared using the single-component systems LNP-53 and LNP-56 respectively, according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 5 μg mRNA / mouse was injected via tail vein. Six hours later, the biofluorescence intensity distribution in mice was measured using an IVIS small animal in vivo imaging system, as well as the biofluorescence intensity distribution in the heart, liver, spleen, lungs, kidneys, and bone marrow after dissection. Mice were intraperitoneally injected with 100 μL of D-luciferin potassium salt (30 mg / mL, dissolved in PBS) before imaging. The results are shown in Figure 9. When using the single-component formulation of Example 15 (LNP to RNA mass ratio of 1.5:1), LNP-luc-mRNA complexes 53 and 56 also showed increased expression in the spleen, consistent with the four-component formulation.

[0438] Example 23: Vaccine prepared from a single-component system leads to activation of antigen-presenting cells in vivo.

[0439] The LNP-OVA-mRNA complex 5 was prepared according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 20 μg mRNA / mouse was injected via tail vein. At 24 h, the activation markers CD40 and CD86 of pDC and cDC in the spleen were detected by flow cytometry. The results are shown in Figures 10-1A and 10-1B. A single intravenous injection of 20 μg of LNP-mRNA encoding OVA induced the maturation of splenic pDCs and cDCs, upregulating the activation markers CD40 and CD86, and simultaneously increasing the proportion of pDCs (Figure 10-2). Serum samples were collected at 6 h and 24 h, and the changes in serum IFNα and IFNy levels were detected by ELISA (Dacowed) . The results are shown in Figure 10-3. Figure 10-3 includes Figures 10-3A and 10-3B. Figure 10-3A shows data 6 hours after injection, and Figure 10-3B shows data 24 hours after injection. Serum IFNα and IFNy levels increased 6 hours after injection and decreased at 24 hours, but remained higher than in wild-type mice. This demonstrates that the vaccine led to the activation of APCs in vivo.

[0440] Example 24: Vaccine prepared in a single-component system leads to antigen-specific T cell activation.

[0441] The LNP-OVA-mRNA complex 5 was prepared according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 20 μg mRNA / mouse was injected via tail vein, and the drug was administered three times at intervals of day 0, day 3, and day 8. On day 13, the level of IFN-γ secreted by the mouse spleen was detected using Elispot (Dakow). The results are shown in Figures 11-1 and 11-2. Unvaccinated mouse spleen cells did not respond to stimulation by the OVA peptide (the amino acid sequence of the OVA peptide SEQ ID NO:1 is SIINFEKL), while vaccinated mice produced T cells that secreted IFN-γ. On day 13, immunophenotyping of the mouse spleen was performed by flow cytometry. The results are shown in Figures 12-1 and 12-2. Vaccination upregulated the proportions of various immune cells, including macrophages, pDCs, cDCs, and CD3+. + T cells, CD8 + T cells, etc. Consistent with the Elispot results, the proportion of CD8+ T cells secreting granulomatin in the spleen was upregulated (Figure 13), indicating that the mRNA vaccine generated a strong T cell response. Serum was collected on day 13, and the vaccine-induced antibody levels were assessed using ELISA (Dacowed). The results are shown in Figures 14-1, 14-2, and 14-3. The proportions of total IgG and IgG2a in the serum increased, demonstrating that the vaccine exerts its immune effect through a synergistic effect of humoral and cellular immunity.

[0442] Example 25: OVA vaccine treatment for solid tumors

[0443] 1. A solid tumor model was established by subcutaneously inoculating B16F10 tumor cells overexpressing OVA antigen to evaluate the anti-solid tumor efficacy of the OVA vaccine. The model mice were randomly divided into three groups: the UT group, the LNP-luc mRNA group, and the LNP-OVA mRNA group, with 10 mice in each group.

[0444] 2. The body weight and tumor size of the mice were measured every two days, and tumor size change curves and survival curves were plotted for the three groups of mice. The tumor size was measured using calipers to measure its length (L) and width (W). The tumor size was calculated using the formula: 1 / 2 × L × W 2 .

[0445] 3. LNP-OVA-mRNA complex 5 was prepared according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 20 μg mRNA / mouse was injected via tail vein, and the vaccine was administered three times at intervals on days 5, 8, and 13 after tumor inoculation. The tumor size change curves (Figure 15) and survival curves (Figure 16) of the two groups of mice demonstrated that the vaccine had good therapeutic efficacy.

[0446] 4. In terms of mouse weight, long-term administration did not cause weight loss; AST detection also proved that LNP did not cause liver damage (as shown in Figures 17-1, 17-2 and 17-3).

[0447] Example 26: Peripheral blood immune cell typing for OVA vaccine treatment of solid tumors

[0448] 1. A solid tumor model was constructed by subcutaneously inoculating B16F10 tumor cells overexpressing OVA antigen. The model mice were randomly divided into three groups: UT group, LNP-luc mRNA group and LNP-OVA mRNA group, with 6 mice in each group.

[0449] 2. Prepare LNP-OVA-mRNA or LNP-luc-mRNA complex 3 according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). Administer 20 μg mRNA / mouse via tail vein injection, and administer three times at intervals on days 5, 8, and 13 after tumor inoculation.

[0450] 3. Blood was collected from the orbital cavity on the 23rd day after tumor grafting. The phenotype of immune cells in the peripheral blood, including immune memory and cytotoxic T cells, was detected by flow cytometry. The results are shown in Figures 18-1, 18-2 and 18-3, which prove that the vaccine can stimulate immune memory and induce T cells to play a killing role.

[0451] Example 27: Tumor microenvironment immune cell typing in OVA vaccine treatment of solid tumors

[0452] 1. The three groups of mice in Example 24 were sacrificed by cervical dislocation on the 24th day after tumor inoculation, and the tumors were cut off;

[0453] 2. Place the tumor to be minced into a 1.5ml EP tube, add 200μl of tissue lysis buffer (Reward), and mince it with scissors until there is basically no obstruction when blowing and aspirating with a 1ml pipette;

[0454] 3. After cutting and adding more than 2 ml of the solution, transfer it to the grinding machine tube using a Pasteur tube. Then add 1 ml of tissue lysis buffer and grind the tissue using the grinding machine.

[0455] 4. After grinding, filter through a 0.45μm filter membrane, transfer to a 15ml centrifuge tube, and centrifuge at 400g for 5min; discard the supernatant, add 6ml of lysing red lysate, and lyse on ice for 5min; then centrifuge at 400g for 5min and discard the supernatant.

[0456] 5. Add 6 ml of DMEM to resuspend, centrifuge at 400g for 5 min, discard the supernatant, resuspend in DMEM medium, and obtain a single-cell suspension;

[0457] 6. The effect of OVA immunotherapy on tumor immune microenvironment reprogramming was investigated by flow cytometry. According to the flow cytometry analysis, compared with the other two groups, the LNP-OVA mRNA group showed an increase in CD45+ immune cells (Figure 19), with an increased proportion of macrophages (Figure 19) and a significant increase in the M1 / M2 ratio (Figure 20). This result demonstrates phenotypic metastasis of pro-tumor macrophages in the tumor microenvironment. Flow cytometry analysis of CD4+ T cells revealed a decrease in Treg cells in the tumor microenvironment of the treatment group (Figure 19), reducing immunosuppression in the tumor microenvironment; while pDC activation indicators showed a significant increase (Figures 21-1 and 21-2), promoting antigen presentation and T cell activation. In conclusion, the OVA vaccine treatment strategy significantly alters the tumor immune microenvironment.

[0458] Example 28: Evaluation of the preventive effect of OVA vaccine

[0459] 1. Wild-type C57 mice were divided into three groups: UT group, LNP-luc mRNA group, and LNP-OVA mRNA group, with 10 mice in each group. LNP-OVA-mRNA or LNP-luc-mRNA complex 3 was prepared according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 20 μg mRNA / mouse was injected via tail vein and administered three times at intervals of day 0, day 3, and day 8.

[0460] 2. On day 15, a solid tumor model was constructed by subcutaneously inoculating B16F10 tumor cells overexpressing OVA antigen to evaluate the preventive effect of the OVA vaccine.

[0461] 3. The weight and tumor size of the mice were measured every two days, and the tumor size change curves (as shown in Figure 22) and survival curves (as shown in Figure 23) of the three groups of mice were plotted. The tumor size was measured using calipers, measuring its length (L) and width (W). The tumor size was calculated using the formula: 1 / 2 × L × W 2 The tumor size change curves and survival curves of the two groups of mice demonstrated that the vaccine has a good preventive effect.

[0462] Example 29: OVA vaccine combined with PD-1 therapy for solid tumors

[0463] 1. A solid tumor model was established by subcutaneously inoculating B16F10 tumor cells overexpressing OVA antigen to evaluate the anti-solid tumor efficacy of OVA vaccine combined with PD-1. The model mice were randomly divided into three groups: UT group, LNP-OVA mRNA group, and LNP-OVA mRNA-PD-1 group, with 10 mice in each group.

[0464] 2. The weight and tumor size of the mice were measured every two days, and tumor size change curves (as shown in Figure 24) and survival curves (as shown in Figure 25) were plotted for the three groups of mice. The tumor size was measured using calipers, measuring its length (L) and width (W). The tumor size was calculated using the formula: 1 / 2 × L × W 2 .

[0465] 3. LNP-OVA-mRNA complex 3 was prepared according to the method in Example 13 (LNP to RNA mass ratio of 1.5:1). 20 μg mRNA / mouse was injected via tail vein, and the drug was administered three times at intervals of day 9, day 13, and day 17 after tumor inoculation. Tumor size change curves and survival curves in both groups of mice demonstrated that the vaccine combined with PD-1 therapy had better therapeutic effects.

[0466] Although the embodiments of this application have been described above in conjunction with the specific embodiments described, this application is not limited to the specific embodiments and application fields described above. The specific embodiments described above are merely illustrative and instructive, and not restrictive. Those skilled in the art can make many other forms based on the teachings of this specification and without departing from the scope of protection of the claims of this application, and these are all within the scope of protection of this application.

Claims

1. Use of a compound of Formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, in targeting immune cells in the spleen, in: G1 and G2 are each independently C2-C24 straight-chain alkylene groups that are optionally substituted with substituents; L1 and L2 are each independently -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-; G3 and G4 are each independently a C2-C24 straight-chain alkylene group, either bonded or optionally substituted with substituents; L3 and L4 are each independently a bond, -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-; G5 and G6 are each independently a C2-C24 straight-chain alkyl group or a C2-C24 straight-chain alkenyl group that is optionally substituted with a substituent. Z is either a C1-C12 alkylene group optionally substituted with a substituent or a -R group optionally substituted with a substituent. e G7R f -; Among them, R e and R f It is a C1-C12 alkylene group that is optionally substituted with a substituent; G7 is -NR g -、-(3-7 membered saturated cycloalkane)-、-(3-7 membered heterocyclic alkane)-、-(3-7 membered cyclic arylene)- or -(3-7 membered cyclic heteroarylene)-; R g Ci-Ci2alkylene that is optionally substituted with a substituent; X and Y are each independently a C1-C12 straight-chain alkyl group optionally substituted with a substituent, or -R k -N(R m (R) n -, -G1L1G3L3G5 or X, Z together with the nitrogen they are attached to form a ring; R k , R m , and R n C1-C12alkyl optionally substituted with substituents; The substituents are selected from one or more of deuterium, tritium, halogen atoms, amino, hydroxyl, cyano, nitro or alkylene groups.

2. Use of a compound of formula (I) or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof in the preparation of a medicament targeting splenic immune cells. in: G1 and G2 are each independently C2-C24 straight-chain alkylene groups that are optionally substituted with substituents; L1 and L2 are each independently -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-; G3 and G4 are each independently a C2-C24 straight-chain alkylene group, either bonded or optionally substituted with substituents; L3 and L4 are each independently a bond, -OC(=O)-, -C(=O)O-, -OC(=O)O-, or -C(=O)-; G5 and G6 are each independently a C2-C24 straight-chain alkyl group or a C2-C24 straight-chain alkenyl group that is optionally substituted with a substituent. Z is either a C1-C12 alkylene group optionally substituted with a substituent or a -R group optionally substituted with a substituent. e G7R f -; Among them, R e and R f It is a C1-C12 alkylene group that is optionally substituted with a substituent; G7 is -NR g -、-(3-7 membered saturated cycloalkane)-、-(3-7 membered heterocyclic alkane)-、-(3-7 membered cyclic arylene)- or -(3-7 membered cyclic heteroarylene)-; R g Ci-Ci2alkylene that is optionally substituted with a substituent; each of said X and Y is independently C1-C12 linear alkyl, -R k -N(R m )(R n )-, -G1L1G3L3G5 or X, Z together with the nitrogen to which they are attached form a ring; R k , R m , and R n C1-C12alkyl optionally substituted with substituents; The substituents are selected from one or more of deuterium, tritium, halogen atoms, amino, hydroxyl, cyano, and nitro groups.

3. Use according to claim 1 or 2, wherein, The compounds have a structure according to Formula (I-A): Wherein, G1, G2, G3, G4, G5, G6, L1, L2, L3, L4, X and Y are as defined in claim 1 or 2; G8 is an optionally substituted C1-C12 alkylene group.

4. Use according to claim 1 or 2, wherein, The compound has the structure shown in formula (IB): Among them, G1, G2, G3, G4, G5, G6, G7, L1, L2, L3, L4, R e R f X and Y are as defined in claim 1 or 2.

5. The use according to claim 1 or 2, wherein, G1 and G2 are unsubstituted C2-C4 alkylene groups.

6. The use according to claim 1 or 2, or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, wherein L1 and L2 are -C(=O)O-.

7. The use according to claim 1 or 2, wherein, G3 and G4 are keys.

8. The use according to claim 1 or 2, or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, wherein G3 and G4 are unsubstituted C2-C4 alkylene groups.

9. The use according to claim 1 or 2, wherein, L3 and L4 are bonds.

10. The use according to claim 1 or 2, or a pharmaceutically acceptable salt, prodrug, or stereoisomer thereof, wherein L3 and L4 are -OC(=O)O-.

11. The use according to claim 1 or 2, wherein, G5 and G6 are optionally substituted C4-C16 straight-chain alkenyl groups.

12. The use according to claim 3, wherein, The compound has the structure shown in formula (IA-1): Where m is 1 or 3, n is 0 or 1, G5 and G6 are defined as in claim 1 or 2, G8 is defined as in claim 3, and G9 and G... 10 For optionally substituted C1-C5 straight-chain alkanes, -R k -N(R m (R) n - or G9 and G 10 Any one of them, along with the nitrogen it is attached to, forms a ring, R K R is an optionally substituted C1-C5 straight-chain alkylene group. m and R n It is a C1-C5 straight-chain alkane that is optionally substituted.

13. The use according to claim 3, wherein, The compounds have a structure according to Formula (I-A-2): G8 is defined as in claim 3, and G5 and G6 are defined as in claim 3.

14. The use according to claim 3, wherein, The compounds have a structure according to Formula (I-A-3): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 3.

15. The use according to claim 3, wherein, The compound has the structure shown in formula (IA-4): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 3.

16. The use of claim 3, wherein, The compound has the structure shown in formula (IA-5): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 3.

17. The use of claim 3, wherein, The compound has a structure shown in formula (I-A-6): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 3.

18. The use of claim 4, wherein, The compound has a structure represented by formula (I-B-1): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 4.

19. The use of claim 4, wherein, The compound has a structure represented by formula (I-B-2): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 4.

20. The use of claim 4, wherein, The compound has a structure represented by formula (I-B-3): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 4.

21. The use of claim 4, wherein, The compound has a structure represented by formula (I-B-4): Where m is 1 or 3, n is 0 or 1, and G5 and G6 are defined as in claim 4.

22. The use according to any one of claims 12-14, wherein, G8 is an optionally substituted C2-C6 alkylene group.

23. The use of claim 12, wherein, G9 or G 10 or both have one of the following structures: methyl, ethyl, 2-hydroxyethyl, -C3H6-N(CH3)(CH3)-.

24. The use according to any one of claims 12-23, wherein, G5 or G6 or both have one of the following structures:

25. The use according to any one of claims 2-24, wherein, The drug that targets spleen immune cells is a vaccine or drug used to treat or prevent tumors, viral infections, or inflammation.

26. A composition wherein, The composition comprises an active component and a carrier for delivering the active component, the active component being a therapeutic or preventative agent; the carrier being a cationic lipid, the cationic lipid being a compound of formula (I) of any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof.

27. The composition of claim 26, wherein, The active component is selected from one or more of nucleic acid molecules, small molecule compounds, polypeptides, or proteins. Preferably, the nucleic acid molecule is selected from one or more of single-stranded DNA, double-stranded DNA, short isomers, agomir, antagomir, antisense molecules, circular RNA, small interfering RNA (siRNA), asymmetric interfering RNA (aiRNA), microRNA (miRNA), Dicersubstrate RNA (dsRNA), small hairpin RNA (shRNA), transfer RNA (tRNA), messenger RNA (mRNA), locked nucleic acid (LNA), peptide nucleic acid (PNA), or morpholine ring oligonucleotides.

28. The composition of claim 26, wherein, The active component comprises at least one mRNA encoding an antigen or a fragment thereof or an epitope, preferably a monocistronic mRNA or a polycistronic mRNA.

29. The composition of claim 28, wherein, The antigens mentioned are tumor-specific antigens or tumor-associated antigens.

30. The composition of claim 28, wherein, The mRNA contains one or more functional nucleotide analogs selected from one or more of pseudouridine, 1-methyl-pseudouridine, or 5-methylcytosine.

31. The compound of claim 27, wherein, The small molecule compound is selected from one or more of the following: antitumor drugs, anti-infective drugs, local anesthetics, antidepressants, anticonvulsants, antibiotics / antibacterial agents, antifungal drugs, antiparasitic drugs, hormones, hormone antagonists, immunomodulators, neurotransmitter antagonists, antiglaucoma agents, anesthetics, or imaging agents.

32. The composition of claim 26, wherein, The mass ratio of the carrier to the active component is 1:10-50:1, preferably 1.5:1.

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