Cationic lipid and use thereof

Abstract

The present disclosure relates to a cationic lipid and use thereof. Specifically provided is a compound represented by formula I or a salt thereof, wherein R1 to R4, H1, H2, and L1 are as defined in the present disclosure. The cationic lipid is used for delivering polynucleotides or nucleic acids and protecting the polynucleotides or nucleic acids from being degraded and removed in serum, and has excellent delivery efficiency.

Description

Cationic lipid and its application Technical Field

[0001] The present invention relates to the field of medicine and a cationic lipid and its application. Background Art

[0002] Gene therapy involves introducing exogenous, normal genes into target cells to correct or compensate for diseases caused by defective or abnormal genes. Since the COVID-19 pandemic, gene therapy has become widely known and has become another hot topic after immunotherapy for cancer, gradually becoming a reality.

[0003] However, naked therapeutic genes are easily degraded by nucleases and exhibit poor cellular uptake. Therefore, the preparation of safe and efficient gene vectors is essential for successful gene therapy. Mainstream nucleic acid drug delivery systems are divided into viral vector systems and non-viral systems, with lipid nanoparticle-mediated nucleic acid drug delivery being the predominant method in non-viral delivery systems.

[0004] Nanoparticles composed of cationic lipids and other auxiliary lipids, such as those formed from cholesterol, phospholipids, and PEGylated lipids, can encapsulate nucleic acids, protecting them from degradation while promoting cellular uptake and reducing immune responses. Furthermore, lipid nanoparticles offer other advantages for cellular delivery of bioactive ingredients, including good targeting, minimal side effects, excellent stability, and high transfection efficiency.

[0005] The rapid development of therapeutic areas based on nucleic acid molecules has placed higher demands on the delivery of nucleic acid drugs, thus creating a need for the development of new cationic lipids and lipid nanoparticles for nucleic acid delivery. Preferably, these cationic lipids provide an excellent drug:lipid ratio, protect nucleic acids from degradation and clearance in serum, are suitable for systemic or local delivery, and provide intracellular delivery of nucleic acids. Summary of the Invention

[0006] The present disclosure provides a compound represented by formula I or a salt thereof

[0007] Among them, H 1 Selected from C 1-12 Alkylene, C 2-12 Alkenylene, C 3-8 Cycloalkylene or C 3-8 cycloalkenylene;

[0008] H 2 Selected from C 1-12 Alkylene or C 1-12 heteroalkylene;

[0009] The condition is that when H 2 C1-12 When alkylene, H 1 Selected from C 3-8 Cycloalkylene or C 3-8 cycloalkenylene;

[0010] R 1 Selected from C 1-24 Alkyl or C 2-24 alkenyl;

[0011] R 2 Selected from C 1-24 Alkyl, C 1-24 Alkenyl or -R b -L 2 -R c ;

[0012] L 1 or L 2 Each independently selected from -C(O)O-, -OC(O)-, -C(O)-, -OC(O)O-, -O-, -S(O)q-, -SS-, -C(O)S-, -SC(O)-, -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a -、-P(O)(OR a ) or key, R a Selected from hydrogen or C 1-6 Alkyl, q is selected from 0, 1 or 2;

[0013] R b Selected from C 1-12 Alkylene or C 2-12 alkenylene;

[0014] R c Selected from C 1-24 Alkyl or C 2-24 alkenyl;

[0015] R 3 Selected from alkylamino or cycloalkylamino;

[0016] R 4 Selected from hydrogen or C 1-6 alkyl.

[0017] In some embodiments, H in the compound of Formula I or its salt is 2 is a heteroalkylene group containing at least one heteroatom selected from O, N and S.

[0018] In some embodiments, H in the compound of Formula I or its salt is2 C 1-12 Heteroalkylene, such as C 2-9 Heteroalkylene, C 2-7 Heteroalkylene, C 2-4 Heteroalkylene, C 3-5 Heteroalkylene or C 3-6 Heteroalkylene.

[0019] In some embodiments, the heteroalkylene group in the compound of Formula I or its salt is a heteroalkylene group containing at least one oxygen atom. In some embodiments, the heteroalkylene group in the compound of Formula I or its salt is a heteroalkylene group containing two oxygen atoms. In some embodiments, the heteroalkylene group in the compound of Formula I or its salt is a heteroalkylene group containing at least one nitrogen atom. In some embodiments, the heteroalkylene group in the compound of Formula I or its salt is a heteroalkylene group containing at least one oxygen atom.

[0020] In some embodiments, R 3 It is an alkylamino group, for example, a methylamino group (CH3NH-) or a dimethylamino group ((CH3)2N-).

[0021] In some embodiments, R 1 C 1-24 Alkyl, including C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, C6 alkyl, C7 alkyl, C8 alkyl, C9 alkyl, C 10 Alkyl, C 11 Alkyl, C 12 Alkyl, C 13 Alkyl, C 14 Alkyl, C 15 Alkyl, C 16 Alkyl, C 17 Alkyl, C 18 Alkyl, C 19 Alkyl, C 20 Alkyl, C 21 Alkyl, C 22 Alkyl or C 23 alkyl.

[0022] In other embodiments, R 1 C 4-18 alkyl.

[0023] In some embodiments, R 1 For straight chain C 4-18 alkyl.

[0024] In some embodiments, R 1 For branched chain C 4-18alkyl.

[0025] In some embodiments, R 1 For branched chain C 17 alkyl.

[0026] In other embodiments, R 1 C 2-24 Alkenyl, including C2 alkenyl, C3 alkenyl, C4 alkenyl, C5 alkenyl, C6 alkenyl, C7 alkenyl, C8 alkenyl, C9 alkenyl, C 10 Alkenyl, C 11 Alkenyl, C 12 Alkenyl, C 13 Alkenyl, C 14 Alkenyl, C 15 Alkenyl, C 16 Alkenyl, C 17 Alkenyl, C 18 Alkenyl, C 19 Alkenyl, C 20 Alkenyl, C 21 In some other embodiments, R 1 C 4-18 Alkenyl.

[0027] In some embodiments, R 1 For straight chain C 4-18 Alkenyl.

[0028] In some embodiments, R 1 For branched chain C 4-18 Alkenyl.

[0029] Other embodiments provide compounds of Formula I or salts thereof wherein R 2 -R b -L 2 -R c , where R b , L 2 or R c As defined above.

[0030] Furthermore, some embodiments provide compounds of formula I as compounds of formula IIa.

[0031] Where X is -N(R d )- or -O-, o is selected from an integer between 1 and 11, p is selected from an integer between 1 and 5, and o+p is not greater than 12, R d Selected from hydrogen or C 1-6 Alkyl, R 5 、R 6、R 7 、R 8 are independently selected from hydrogen, halogen, hydroxyl or C 1-6 Alkyl, L 1 , L 2 、H 1 、R 1 、R 3 、R 4 、R b and R c As defined in the compound represented by formula I.

[0032] In some embodiments, R b C 1-12 Alkylene, including C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C 12 Alkylene.

[0033] In some embodiments, R b For straight chain C 3-12 Alkylene.

[0034] In some embodiments, R b For branched chain C 3-12 Alkylene.

[0035] In some embodiments, R b It is a straight chain C7 alkylene group.

[0036] In some embodiments, R b It is a straight chain C8 alkylene group.

[0037] In some embodiments, R b It is a straight chain C9 alkylene group.

[0038] On the other hand, in the compound represented by Formula I or IIa or its salt, R b C 1-12 Alkenylene, including C3 alkenylene, C4 alkenylene, C5 alkenylene, C6 alkenylene, C7 alkenylene, C8 alkenylene, C9 alkenylene, C 10 Alkenylene, C 11 Alkenylene or C 12 Alkenylene.

[0039] In some embodiments, Rb For branched chain C 3-12 Alkenylene.

[0040] In some embodiments, R b It is a straight chain C7 alkenylene.

[0041] In some embodiments, R b It is a straight-chain C8 alkenylene group.

[0042] In some embodiments, R b It is a straight-chain C9 alkenylene group.

[0043] In other embodiments, R c C 1-24 Alkyl, including C2 alkyl, C4 alkyl, C5 alkyl, C6 alkyl, C7 alkyl, C8 alkyl, C9 alkyl, C 10 Alkyl, C 11 Alkyl, C 12 Alkyl, C 13 Alkyl, C 14 Alkyl, C 15 Alkyl, C 16 Alkyl, C 17 Alkyl, C 18 Alkyl, C 19 Alkyl, C 20 Alkyl, C 21 Alkyl, C 22 Alkyl, C 23 Alkyl or C 24 alkyl.

[0044] In some embodiments, R c For straight chain C 1-12 alkyl.

[0045] In some embodiments, R c For branched chain C 1-12 alkyl.

[0046] In some embodiments, R c For branched chain C 11 alkyl.

[0047] In some embodiments, R c For branched chain C 17 alkyl.

[0048] In some embodiments, R c It is a straight chain C9 alkyl group.

[0049] In other embodiments, R c C 2-24 Alkenyl, including C2 alkenyl, C4 alkenyl, C5 alkenyl, C6 alkenyl, C7 alkenyl, C8 alkenyl, C9 alkenyl, C 10 Alkenyl, C 11 Alkenyl, C 12 Alkenyl, C 13 Alkenyl, C 14 Alkenyl, C 15 Alkenyl, C 16 Alkenyl, C 17 Alkenyl, C 18 Alkenyl, C 19 Alkenyl, C 21 Alkenyl, C 22 Alkenyl, C 23 Alkenyl or C 24 Alkenyl.

[0050] In some embodiments, R c For straight chain C 1-12 Alkenyl.

[0051] In some embodiments, R c For branched chain C 1-12 Alkenyl.

[0052] In some embodiments, R c For branched chain C 11 Alkenyl.

[0053] In some embodiments, R c For branched chain C 17 Alkenyl.

[0054] In some embodiments, R c It is a straight chain C9 alkenyl.

[0055] In some embodiments, p in the compound represented by Formula I or IIa or a salt thereof is selected from 1 or 2.

[0056] In some embodiments, o in the compound represented by Formula I or IIa or a salt thereof is selected from an integer between 4 and 7, such as 5 or 6.

[0057] In some embodiments, X in the compound of Formula I or IIa or a salt thereof is -O-.

[0058] In some embodiments, in the compound represented by Formula I or IIa or a salt thereof, p is selected from 1 or 2; and X is -O-.

[0059] In some embodiments, in the compound represented by Formula I or II or a salt thereof, p is selected from 3 or 4, o is selected from 5 or 6, and X is -O-.

[0060] In some embodiments, R 5 、R 6 Each independently is C 1- 6-alkyl, such as methyl, ethyl or propyl.

[0061] In some embodiments, R 7 、R 8 Each independently is C 1- 6-alkyl, such as methyl, ethyl or propyl.

[0062] In some embodiments, R 5 、R 6 、R 7 、R 8 are each independently hydrogen.

[0063] In some embodiments, in the compound represented by Formula I or II or a salt thereof, p is selected from 3 or 4, o is selected from 5 or 6, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen.

[0064] In some embodiments, L in the compound represented by Formula I or IIa or a salt thereof 1 and L 2 are each independently selected from -C(O)O-, -OC(O)- or a bond.

[0065] In some embodiments, R 4 C 1-6 Alkyl groups, such as methyl or ethyl.

[0066] In some embodiments, R 4 For hydrogen.

[0067] In some embodiments, in the compound of formula I or II or its salt, p is selected from 3 or 4, o is selected from 5 or 6, X is -O-, R 5 、R 6 、R 7 、R 8are each independently hydrogen, R 4 For hydrogen.

[0068] In some embodiments, in the compound of formula I or II or a salt thereof, p is selected from 1 or 2, o is selected from 5 or 6, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen, R 4 In some embodiments, in the compound represented by Formula I or II or its salt, p is selected from 1 or 2, o is selected from 5 or 6, X is -O-, R 5 、R 6 、R 7 、R 8 are each independently hydrogen, R 4 is hydrogen, L 1 and L 2 Each is independently selected from -C(O)O- or -OC(O)-.

[0069] In some embodiments, in the compound represented by Formula I or II or a salt thereof, o is selected from 1 or 2, p is selected from 4 or 5, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen, R 4 For hydrogen.

[0070] In some embodiments, in the compound represented by Formula I or II or a salt thereof, o is selected from 1 or 2, p is selected from 4 or 5, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen, R 4 is hydrogen, L 1 and L 2 Each is independently selected from -C(O)O- or -OC(O)-.

[0071] In some embodiments, the compound represented by formula IIa is a compound represented by formula IIa-1 or a compound represented by formula IIa-2

[0072] In some of the embodiments, in the compound represented by formula IIa-1 or IIa-1 or its salt, o is selected from 1 or 2, p is selected from 4 or 5, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen.

[0073] In some of the embodiments, in the compound represented by formula IIa-1 or IIa-1 or its salt, p is selected from 1 or 2, o is selected from 4 or 5, X is -O-, and R 5 、R 6 、R 7 、R 8 are each independently hydrogen.

[0074] On the other hand, in certain embodiments, L in the compound represented by Formula I or IIa or its salt is 1 and L 2 are each independently selected from -C(O)-, -OC(O)O-, -O- or a bond.

[0075] In certain embodiments, L in the compound represented by Formula I or IIa or its salt 1 and L 2 Each independently selected from -S(O) q -, -SS-, -C(O)S-, -SC(O)- or a bond.

[0076] In certain embodiments, L in the compound represented by Formula I or IIa or its salt 1 and L 2 Each independently selected from -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O- or bond.

[0077] In certain embodiments, L in the compound represented by Formula I or IIa or its salt 1 and L 2 Each independently selected from -NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a - or key.

[0078] In some embodiments, H in the compound represented by Formula I or IIa is 1 Selected from C 1-12 Alkylene or C 2-12 Alkenylene.

[0079] In some embodiments, H in the compound represented by Formula I or IIa is 1 C 1-12 Alkylene, including C2 alkylene, C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C12 Alkylene.

[0080] In some embodiments, H in the compound represented by Formula I or IIa is 1 It is a C3 alkylene group, for example a straight-chain C3 alkylene group.

[0081] In some embodiments, H in the compound represented by Formula I or IIa is 1 It is a C4 alkylene group, for example a straight-chain C3 alkylene group.

[0082] In some embodiments, H in the compound represented by Formula I or IIa is 1 It is a C5 alkylene group, for example a linear C3 alkylene group.

[0083] In some embodiments, in the compound of formula I or II or its salt, p is selected from 3 or 4, o is selected from 5 or 6, X is -O-, R 5 、R 6 、R 7 、R 8 are each independently hydrogen, H 1 is a straight chain C3 alkylene, R b For straight chain C 6- 8 alkylene.

[0084] On the other hand, H in the compound represented by formula I 1 C 3-8 Cycloalkylene, for example cyclohexylene.

[0085] In some embodiments, the compound represented by formula IIa is a compound represented by formula IIa-1a or a compound represented by formula IIa-2a

[0086] In some embodiments, R in the compound represented by formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt 7 、R 8 are each independently hydrogen.

[0087] In some embodiments, o in the compound represented by Formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt is selected from 5 or 6.

[0088] In some embodiments, R in the compound represented by formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt 5 、R 6 are each independently hydrogen.

[0089] In some embodiments, H in the compound represented by Formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt 1 It is a C3 alkylene group, for example a straight-chain C3 alkylene group.

[0090] In some embodiments, H in the compound represented by Formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt 1 It is a C3 alkylene group, for example a straight-chain C3 alkylene group.

[0091] In some embodiments, R in the compound represented by formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt b For branched chain C 3-12 Alkylene.

[0092] In some embodiments, R in the compound represented by formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt b It is a straight chain C7 alkylene group.

[0093] In some embodiments, R in the compound represented by formula IIa-1 or IIa-2 or IIa-1a or IIa-2a or its salt c for

[0094] In some embodiments, the compound represented by formula IIa or IIa-2 or its salt is a compound represented by formula IIa-3 or its salt

[0095] wherein o and p are independently selected from integers between 1 and 6, and o+p=6 or 7, and X is -N(R d )-or-O-, R d Selected from hydrogen or C 1-6 alkyl.

[0096] In some embodiments, in the compound represented by Formula IIa-3 or a salt thereof, o is selected from 1 or 2, p is selected from 4 or 5, and X is -O-.

[0097] In some embodiments, in the compound represented by Formula IIa-3 or a salt thereof, p is selected from 1 or 2, o is selected from 4 or 5, and X is -O-.

[0098] In some embodiments, the compound of formula I or its salt is not

[0099] Where o=3, p=3.

[0100] In some embodiments, the compound represented by formula I is a compound represented by formula IIb

[0101] Among them L 1 , L 2 、H 2 、R 1 、R 3 、R4 、R b and R c As defined in the compound represented by formula I.

[0102] In other embodiments, L in the compound represented by Formula I or IIb or its salt 1 and L 2 are each independently selected from -OC(O)O- or a bond.

[0103] In other embodiments, R 4 For hydrogen.

[0104] In some embodiments, the compound represented by formula IIb is a compound represented by formula IIb-1 or a compound represented by formula IIb-2

[0105] In certain embodiments, L in the compound represented by Formula I or IIb or its salt 1 and L 2 are each independently selected from -C(O)-, -OC(O)O-, -O- or a bond.

[0106] In certain embodiments, L in the compound represented by Formula I or IIb or its salt 1 and L 2 Each independently selected from -S(O) q -, -SS-, -C(O)S-, -SC(O)- or a bond.

[0107] In certain embodiments, L in the compound represented by Formula I or IIb or its salt 1 and L 2 Each independently selected from -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O- or bond.

[0108] In certain embodiments, L in the compound represented by Formula I or IIb or its salt 1 and L 2 Each independently selected from -NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a - or key.

[0109] In other embodiments, H in the compound represented by Formula I or IIb or its salt 2 C 1-12Alkylene, including C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C 12 Alkylene.

[0110] In some embodiments, H in the compound represented by Formula I or IIb or a salt thereof 2 For straight chain C 3-12 Alkylene.

[0111] In some embodiments, H in the compound represented by Formula I or IIb or a salt thereof 2 For branched chain C 3-12 Alkylene.

[0112] In some embodiments, H in the compound represented by Formula I or IIb or a salt thereof 2 It is a straight chain C7 alkylene group.

[0113] In some embodiments, H in the compound represented by Formula I or IIb or a salt thereof 2 It is a straight chain C8 alkylene group.

[0114] In some embodiments, H in the compound represented by Formula I or IIb or a salt thereof 2 It is a straight chain C9 alkylene group.

[0115] On the other hand, in some embodiments, H in the compound represented by Formula IIb or its salt is 2 C 1-12 Heteroalkylene, such as C 2-9 Heteroalkylene, C 2-7 Heteroalkylene, C 2-4 Heteroalkylene, C 3-5 Heteroalkylene or C 3-6 Heteroalkylene.

[0116] In some embodiments, the heteroalkylene group in the compound of formula IIb or its salt is a heteroalkylene group containing at least one oxygen atom. In some embodiments, the heteroalkylene group in the compound of formula IIb or its salt is a heteroalkylene group containing two oxygen atoms. In some embodiments, the heteroalkylene group in the compound of formula IIb or its salt is a heteroalkylene group containing at least one nitrogen atom. In some embodiments, the heteroalkylene group in the compound of formula IIb or its salt is a heteroalkylene group containing at least one oxygen atom.

[0117] In certain embodiments, R b For straight chain C 3-12 Alkylene, H 2 C 1-12 Heteroalkylene, such as C7 alkylene or C8 alkylene.

[0118] In certain embodiments, R b For straight chain C 3-12 Alkylene, H 2 C 1-12 Alkylene, for example C7 alkylene or C8 alkylene.

[0119] Some embodiments provide compounds of formula I wherein R c Selected from:

[0120] On the other hand, some embodiments provide compounds of Formula I wherein R 1 Selected from:

[0121] Typical compounds shown in Formula I include:

[0122] On the other hand, the present disclosure also provides a compound represented by formula A or a salt thereof,

[0123] Among them, M 1 Each independently selected from C 1-12 Alkylene, C 2-12 Alkenylene, C 3-8 Cycloalkylene or C 3-8 cycloalkenylene;

[0124] M 2 Each independently selected from C 1-12 Alkylene or C 1-12 heteroalkylene;

[0125] R 1 Each independently selected from C 1-24 Alkyl or C 2-24 alkenyl;

[0126] R 2 Each independently selected from C 1-24 Alkyl, C 1-24 Alkenyl or -R b -L 2 -R c ;

[0127] L 1 or L 2 Each independently selected from -C(O)O-, -OC(O)-, -C(O)-, -OC(O)O-, -O-, -S(O)q-, -SS-, -C(O)S-, -SC(O)-, -NR a C(O)-、-C(O)NR a -、-NRa C(O)NR a -、-NR a C(O)O-、-OC(O)NR a -、-P(O)(OR a ) or key, R a Selected from hydrogen or C 1-6 Alkyl or C 2-6 alkenyl, q is selected from 0, 1 or 2;

[0128] R b Each independently selected from C 1-12 Alkylene or C 2-12 alkenylene;

[0129] R c Each independently selected from C 1-24 Alkyl or C 2-24 alkenyl;

[0130] R 4 are each independently selected from hydrogen or C 1-6 alkyl.

[0131] In some embodiments, M in the compound represented by Formula A or its salt is 2 Each independently is C 1-12 Alkylene, including C2 alkylene, C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C 12 Alkylene.

[0132] In some embodiments, M in the compound represented by Formula A or its salt is 2 Each is independently a C7 alkylene group, for example a linear C7 alkylene group.

[0133] In some embodiments, M in the compound represented by Formula A or its salt is 2 Each is independently a C8 alkylene group, for example a linear C8 alkylene group.

[0134] In other embodiments, M in the compound represented by formula A or its salt 2 are each independently a heteroalkylene group containing at least one heteroatom selected from O, N, and S.

[0135] In certain embodiments, the compound represented by formula A or its salt is 2 Each independently is C 1-12 Heteroalkylene, such as C 2-9 Heteroalkylene, C 2-7 Heteroalkylene, C 2-4 Heteroalkylene, C 3-5Heteroalkylene or C 3-6 Heteroalkylene.

[0136] On the other hand, in some embodiments, M in the compound represented by formula A is 1 Each independently selected from C 1-12 Alkylene or C 2-12 Alkenylene.

[0137] In some embodiments, in the compound represented by formula A, M 1 Each independently is C 1-12 Alkylene, including C2 alkylene, C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C 12 Alkylene.

[0138] In some embodiments, in the compound represented by formula A, M 1 Each is independently a C3 alkylene group, for example a linear C3 alkylene group.

[0139] In some embodiments, in the compound represented by formula A, M 1 Each is independently a C4 alkylene group, such as a linear C3 alkylene group.

[0140] In some embodiments, in the compound represented by formula A, M 1 Each is independently a C5 alkylene group, for example a linear C3 alkylene group.

[0141] Other embodiments provide compounds represented by formula A or salts thereof wherein R 2 -R b -L 2 -R c , where R b , L 2 and R c As defined above.

[0142] In some embodiments, L in the compound represented by Formula A or its salt 1 and L 2 are each independently selected from -C(O)O-, -OC(O)-, and a bond.

[0143] In some embodiments, the compound represented by formula A is a compound represented by formula Aa-1 or formula Aa-2

[0144] In certain embodiments, L in the compound represented by formula A or its salt 1 and L 2 are each independently selected from -C(O)-, -OC(O)O-, -O- or a bond.

[0145] In certain embodiments, L in the compound represented by formula A or its salt 1 and L 2 Each independently selected from -S(O) q -, -SS-, -C(O)S-, -SC(O)- or a bond.

[0146] In certain embodiments, L in the compound represented by formula A or its salt 1 and L 2 Each independently selected from -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O- or bond.

[0147] In certain embodiments, L in the compound represented by formula A or its salt 1 and L 2 Each independently selected from -NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a - or key.

[0148] On the other hand, in the compound represented by formula A, M 1 Each independently is C 3-8 Cycloalkylene, for example cyclohexylene.

[0149] In some embodiments, the compound represented by formula A is a compound represented by formula Ab-1 or Ab-2

[0150] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 Each independently is C 1-24 Alkyl, including C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, C6 alkyl, C7 alkyl, C8 alkyl, C9 alkyl, C 10 Alkyl, C 11 Alkyl, C 12 Alkyl, C 13 Alkyl, C 14 Alkyl, C 15 Alkyl, C 16 Alkyl, C 17 Alkyl, C 18 Alkyl, C 19 Alkyl, C 20 Alkyl, C 21 Alkyl, C 22 Alkyl or C23 alkyl.

[0151] In other embodiments, R 1 Each independently is C 4-18 alkyl.

[0152] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 For straight chain C 4-18 alkyl.

[0153] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 For branched chain C 4-18 alkyl.

[0154] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 For branched chain C 17 alkyl.

[0155] In other embodiments, R 1 Each independently is C 2-24 Alkenyl, including C2 alkenyl, C3 alkenyl, C4 alkenyl, C5 alkenyl, C6 alkenyl, C7 alkenyl, C8 alkenyl, C9 alkenyl, C 10 Alkenyl, C 11 Alkenyl, C 12 Alkenyl, C 13 Alkenyl, C 14 Alkenyl, C 15 Alkenyl, C 16 Alkenyl, C 17 Alkenyl, C 18 Alkenyl, C 19 Alkenyl, C 20 Alkenyl, C 21 Alkenyl.

[0156] In some other embodiments, the compound of formula I or its salt is provided as follows: 1 C 4-18 Alkenyl.

[0157] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 For straight chain C 4-18 Alkenyl.

[0158] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 1 For branched chain C 4-18 Alkenyl.

[0159] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b Each independently is C 1-12 Alkylene, including C3 alkylene, C4 alkylene, C5 alkylene, C6 alkylene, C7 alkylene, C8 alkylene, C9 alkylene, C 10 Alkylene, C 11 Alkylene or C 12 Alkylene.

[0160] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b For straight chain C 3-12 Alkylene.

[0161] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b For branched chain C 3-12 Alkylene.

[0162] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight chain C7 alkylene group.

[0163] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight chain C8 alkylene group.

[0164] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight chain C9 alkylene group.

[0165] On the other hand, in the compound represented by formula A, formula Aa-1, formula Aa-2, formula Ab-1, formula Ab-2, or a salt thereof, R b Each independently is C 2-12 Alkenylene, including C3 alkenylene, C4 alkenylene, C5 alkenylene, C6 alkenylene, C7 alkenylene, C8 alkenylene, C9 alkenylene, C 10 Alkenylene, C 11 Alkenylene or C 12 Alkenylene.

[0166] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b For branched chain C 3-12 Alkenylene.

[0167] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight chain C7 alkenylene.

[0168] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight-chain C8 alkenylene group.

[0169] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is b It is a straight-chain C9 alkenylene group.

[0170] In other embodiments, R c Each independently is C 1-24 Alkyl, including C2 alkyl, C4 alkyl, C5 alkyl, C6 alkyl, C7 alkyl, C8 alkyl, C9 alkyl, C 10 Alkyl, C 11 Alkyl, C 12 Alkyl, C 13 Alkyl, C 14 Alkyl, C 15 Alkyl, C 16 Alkyl, C 17 Alkyl, C 18 Alkyl, C 19 Alkyl, C 20 Alkyl, C 21 Alkyl, C 22 Alkyl, C 23 Alkyl or C 24 alkyl.

[0171] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For straight chain C 1-12 alkyl.

[0172] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For branched chain C 1-12 alkyl.

[0173] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For branched chain C 11 alkyl.

[0174] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For branched chain C 17 alkyl.

[0175] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c It is a straight chain C9 alkyl group.

[0176] In other embodiments, R c Each independently is C 2-20 Alkenyl, including C2 alkenyl, C4 alkenyl, C5 alkenyl, C6 alkenyl, C7 alkenyl, C8 alkenyl, C9 alkenyl, C 10 Alkenyl, C 11 Alkenyl, C 12 Alkenyl, C 13 Alkenyl, C 14 Alkenyl, C 15 Alkenyl, C 16 Alkenyl, C 17 Alkenyl, C 18 Alkenyl, C 19 Alkenyl or C 20 Alkenyl.

[0177] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For straight chain C 1-12 Alkenyl.

[0178] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For branched chain C 1-12 Alkenyl.

[0179] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c For branched chain C 11 Alkenyl.

[0180] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is cFor branched chain C 17 Alkenyl.

[0181] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is c It is a straight chain C9 alkenyl.

[0182] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 4 are each independently hydrogen.

[0183] In some embodiments, R in the compound represented by Formula A or Formula Aa-1 or Formula Aa-2 or Formula Ab-1 or Formula Ab-2 or its salt is 4 Each independently is C 1-6 Alkyl groups, such as methyl or ethyl.

[0184] On the other hand, some embodiments provide compounds represented by formula A or formula Aa-1 or formula Aa-2 or formula Ab-1 or formula Ab-2 wherein R 1 Selected from:

[0185] Typical compounds shown in formula A include:

[0186] The present disclosure also provides an isotope substitution of the aforementioned compound or a salt thereof. In some embodiments, the isotope substitution is a deuterium atom substitution.

[0187] The present disclosure also provides a lipid particle comprising the aforementioned compound or a salt thereof or an isotope thereof. Furthermore, in some embodiments, the lipid particle further comprises an active agent.

[0188] In some embodiments, the active agent is selected from a polynucleotide or a nucleic acid (such as ribonucleic acid or deoxyribonucleic acid). In some embodiments, the active agent is selected from mRNA.

[0189] The present disclosure also provides a pharmaceutical composition comprising the aforementioned lipid particles and a pharmaceutically acceptable excipient. In certain embodiments, based on the total weight of the composition, the pharmaceutical composition contains 0.01%-99.99% of a pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition contains 0.1%-99.9% of a pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition contains 0.5%-99.5% of a pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition contains 1%-99% of a pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition contains 2%-98% of a pharmaceutically acceptable excipient.

[0190] In some embodiments, the excipients include but are not limited to buffers and lyoprotectants.

[0191] In some embodiments, the pharmaceutical composition contains a buffer, such as Tris hydrochloride.

[0192] In some embodiments, the pharmaceutical composition contains a lyoprotectant, such as sucrose.

[0193] The present disclosure also provides a use of the aforementioned compound or its salt, or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition in the preparation of a drug for inducing an immune response in a subject.

[0194] The present disclosure also provides use of the aforementioned compound or its salt, or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition in preparing a medicament for preventing and / or treating diseases or conditions associated with overexpression of the polypeptide.

[0195] The present disclosure also provides use of the aforementioned compound or its salt, or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition in the preparation of a medicament for preventing and / or treating diseases or conditions associated with insufficient polypeptide expression.

[0196] In some embodiments, the disease or condition includes, but is not limited to, cancer, infection, autoimmune disease, neurodegenerative disease, and inflammation.

[0197] The present disclosure also provides a method for preventing and / or treating a disease or condition that induces an immune response in a subject, comprising administering to the patient a composition containing the aforementioned compound or its salt or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition.

[0198] The present disclosure also provides a method for preventing and / or treating a disease or condition associated with overexpression of a polypeptide, comprising administering to the patient a composition containing the aforementioned compound or a salt or isotope thereof, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition.

[0199] The present disclosure also provides a method for preventing and / or treating a disease or condition associated with insufficient polypeptide expression, comprising administering to the patient a composition containing the aforementioned compound or a salt or isotope thereof, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition.

[0200] On the other hand, the present disclosure further provides the aforementioned compound or its salt or isotope substitute, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition, for use in preventing and / or treating a disease or condition that induces an immune response in a subject.

[0201] The present disclosure also provides the aforementioned compound or its salt or isotope substitute, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition, for use in preventing and / or treating diseases or disorders associated with overexpression of the polypeptide.

[0202] The present disclosure also provides the aforementioned compound or its salt or isotope substitute, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition, for use in preventing and / or treating diseases or conditions associated with insufficient polypeptide expression.

[0203] The present disclosure also provides use of the aforementioned compound or its salt, or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition in the preparation of a medicament for preventing and / or treating cancer, infection, autoimmune disease, neurodegenerative disease and inflammation.

[0204] The present disclosure also provides a method for preventing and / or treating cancer, infection, autoimmune disease, neurodegenerative disease and inflammation, comprising administering to the patient a composition containing the aforementioned compound or its salt or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition.

[0205] The present disclosure also provides the aforementioned compound or its salt or isotope substitution, or the aforementioned lipid particle, or the aforementioned pharmaceutical composition, for use in preventing and / or treating cancer, infection, autoimmune disease, neurodegenerative disease and inflammation.

[0206] The compound salts described in the present disclosure include "acid" addition salts and "base" addition salts. For example, salts formed by acid-base reaction with a basic group (amino group), wherein the acid includes an organic acid or an inorganic acid. In addition, the compound salts also include salts formed by quaternization with a basic group (amino group), wherein the quaternizing agent includes a linear or branched chlorinated hydrocarbon.

[0207] The compounds of the present disclosure may exist in specific geometric or stereoisomeric forms. The present disclosure contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present disclosure. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All of these isomers and their mixtures are included within the scope of the present disclosure. The compounds of the present disclosure containing asymmetric carbon atoms can be isolated in optically pure form or in racemic form. Optically pure forms can be resolved from racemic mixtures or synthesized by using chiral starting materials or chiral reagents.

[0208] Optically active (R)- and (S)-isomers, as well as D and L isomers, can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the present disclosure is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting diastereomeric mixture is separated and the auxiliary group is cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), a diastereomeric salt is formed with an appropriate optically active acid or base, and then the diastereoisomers are separated by conventional methods known in the art, and then the pure enantiomer is recovered. In addition, the separation of enantiomers and diastereomers is typically accomplished using chromatography, which employs a chiral stationary phase and is optionally combined with a chemical derivatization method (e.g., carbamate formation from an amine).

[0209] In the chemical structures of the compounds disclosed herein, the bond Indicates that the configuration is not specified, that is, if chiral isomers exist in the chemical structure, the bond Can be or include both Two configurations. In the chemical structure of the compound disclosed in the present invention, the bond The configuration is not specified, that is, it can be Z configuration or E configuration, or contain both configurations.

[0210] Compounds and intermediates of the present disclosure can also exist in different tautomeric forms, and all such forms are included in the scope of the present disclosure. The term "tautomer" or "tautomeric form" refers to structural isomers of different energies that can interconvert via a low energy barrier. For example, proton tautomers (also referred to as prototransfer tautomers) include interconversions via proton migration, such as keto-enol and imine-enamine, lactam-lactim isomerization. The lactam-lactim equilibrium example is between A and B as shown below.

[0211] All compounds in this disclosure can be drawn as either Form A or Form B. All tautomeric forms are within the scope of this disclosure. The naming of the compounds does not exclude any tautomers.

[0212] The present disclosure also includes isotopically labeled compounds of the present disclosure that are identical to those described herein, but where one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into the compounds of the present disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as 2 H. 3 H. 11 C. 13 C.14 C. 13 N. 15 N. 15 O. 17 O. 18 O. 31 P. 32 P. 35 S. 18 F. 123 I. 125 I and 36 Cl et al.

[0213] Unless otherwise stated, when a position is specifically designated as deuterium (D), the position is understood to have at least 1000 times the abundance of deuterium greater than the natural abundance of deuterium (which is 0.015%) (i.e., at least 10% deuterium incorporation). In the example, the compound has a natural abundance greater than deuterium that can be at least 1000 times the abundance of deuterium, at least 2000 times the abundance of deuterium, at least 3000 times the abundance of deuterium, at least 4000 times the abundance of deuterium, at least 5000 times the abundance of deuterium, at least 6000 times the abundance of deuterium or more abundant deuterium. The disclosure also includes various deuterated forms of formula (I) compounds. Each available hydrogen atom connected to a carbon atom can be independently replaced by a deuterium atom. Those skilled in the art can synthesize deuterated forms of formula (I) compounds with reference to relevant literature. Commercially available deuterated starting materials may be used in the preparation of deuterated forms of the compounds of formula (I), or they may be synthesized using conventional techniques using deuterated reagents, including but not limited to deuterated borane, trideuterated borane in tetrahydrofuran, deuterated lithium aluminum hydride, deuterated iodoethane, deuterated iodomethane, and the like.

[0214] "Optionally" or "optionally" means that the subsequently described event or circumstance may but need not occur, and the description includes instances where the event or circumstance occurs or does not occur. For example, "optionally substituted C 1- The term "alkyl" means that halogen or cyano may but need not be present, and the description includes the case where the alkyl is substituted by halogen or cyano and the case where the alkyl is not substituted by halogen and cyano.

[0215] Explanation of terms:

[0216] A "pharmaceutical composition" refers to a mixture containing one or more compounds described herein, or their physiologically acceptable salts or prodrugs, together with other chemical components, as well as other components such as physiologically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitating absorption of the active ingredients and thereby exerting their biological activity.

[0217] "Pharmaceutically acceptable excipients" or "pharmaceutically acceptable excipients" include, but are not limited to, any adjuvant, carrier, glidant, sweetener, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, solvent or emulsifier that has been approved by the U.S. Food and Drug Administration for use in humans or domestic animals.

[0218] "Buffer" refers to a buffer that resists changes in pH through the action of its acid-base conjugate components. Examples of buffers that control pH in an appropriate range include acetate or tris hydrochloride (Tris hydrochloride).

[0219] As used herein, an "effective amount," "therapeutically effective amount," or "therapeutically effective amount" encompasses an amount sufficient to ameliorate or prevent the symptoms or conditions of a medical condition. An effective amount also refers to an amount sufficient to permit or facilitate diagnosis. The effective amount for a particular patient or veterinary subject may vary depending on factors such as the condition to be treated, the patient's overall health, the route and dosage of administration, and the severity of side effects. An effective amount can be the maximum dose or dosage regimen that avoids significant side effects or toxic effects.

[0220] The term "nucleic acid" is a polymer composed of nucleotides, such as deoxyribonucleotides (DNA) or ribonucleotides (RNA).

[0221] The term "oligonucleotide" refers to a single-stranded or double-stranded nucleotide polymer of 2 to 100 nucleotides in length. "Polynucleotide" refers to a single-stranded or double-stranded polymer composed of nucleotide monomers. In some embodiments, a polynucleotide is composed of more than 100 nucleotides in length.

[0222] In other embodiments, exemplary polynucleotides include, but are not limited to, deoxyribonucleotides (DNA), ribonucleic acids (RNA), including messenger RNA, RNAi-inducing agents, shRNA, siRNA, miRNA, antisense RNA, and the like.

[0223] The terms "polypeptide," "peptide," and "protein" are used interchangeably herein to refer to polymers of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residues is an artificial chemical analogue of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers.

[0224] "Alkyl" refers to a saturated aliphatic hydrocarbon group, including straight-chain and branched-chain alkyl groups. In some embodiments, the alkyl group has 1-24 carbon atoms, also known as C 1-24 In some embodiments, an alkyl group has 2-24 carbon atoms, also known as a C 2-24In some embodiments, the alkyl group has 4-18 carbon atoms, also known as C 4-18 In some embodiments, an alkyl group has 1-12 carbon atoms, also known as a C 1-12 In some embodiments, an alkyl group has 1-4 carbon atoms, also known as a C 1-4 In some embodiments, an alkyl group has 10-22 carbon atoms, also known as a C 10-22 In some embodiments, the alkyl group has 4-22 carbon atoms, also known as C 4-22 alkyl.

[0225] The alkyl group may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the alkyl group is substituted with one or more alkyl groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0226] "Alkoxy" refers to -O-(alkyl), where alkyl is as defined above. Non-limiting examples of alkoxy include methoxy and ethoxy. Alkoxy may be unsubstituted or substituted with one or more radicals selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the alkoxy group is substituted by one or more halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0227] "Alkylamino" refers to -NH(alkyl) or -N(alkyl)2, where alkyl is as defined above. Exemplary alkylamino groups include C 1-6 Alkane NH- or di(C 1-6 Alkyl) N-, for example methylamino or dimethylamino. Alkylamino may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the alkylamino group is substituted by one or more alkylamino groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0228] "Heteroalkyl" refers to a straight or branched chain alkyl group preferably having 1 to 14 carbons, more preferably 1 to 12 carbons, even more preferably 2 to 10 carbons in the chain, wherein one or more carbons are substituted with a heteroatom selected from S, O, and N, including -OO- or -SS- moieties. Exemplary heteroalkyl groups include wait.

[0229] Heteroalkyl may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the heteroalkyl group is substituted with one or more halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0230] "Alkenyl" refers to an unsaturated aliphatic hydrocarbon group, including straight-chain and branched alkenyl groups. In some embodiments, the alkenyl group has 2-24 carbon atoms, also known as C 2-24 In some embodiments, the alkenyl group has 2-12 carbon atoms and is also referred to as a C 2-12 In some embodiments, the alkenyl group has 4-18 carbon atoms and is also referred to as a C 4-18 In some embodiments, the alkenyl group has 1-4 carbon atoms, also known as C 1-4 In some embodiments, the alkenyl group has 10-22 carbon atoms and is also referred to as a C 10-22 In some embodiments, the alkenyl group has 4-22 carbon atoms and is also referred to as a C 4-22 Alkenyl. Exemplary alkenyl groups include ethenyl, propenyl, n-butenyl, isobutenyl, 3-methylbut-2-enyl, n-pentenyl, heptenyl, octenyl, cyclohexyl-butenyl, and decenyl.

[0231] Alkenyl groups may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the alkenyl group is substituted with one or more halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0232] "Heteroalkenyl" refers to the definition of "heteroalkyl". It refers to a straight chain or branched alkenyl group in which one or more carbon atoms are replaced by heteroatoms selected from S, O and N, including -OO- or -SS- moieties.

[0233] A "monovalent group" is a compound formed by formally eliminating a monovalent atom or group. A "subunit" is a compound formed by formally eliminating two monovalent or one divalent atom or group.

[0234] The term "alkylene" refers to the portion of an alkane molecule remaining after removing two hydrogen atoms. In some embodiments, an alkylene group has 1-12 carbon atoms and is also referred to as a C 1-12 In some embodiments, an alkylene group has 3-8 carbon atoms, also known as a C 3-8 In some embodiments, an alkylene group has 1-4 carbon atoms, also known as C 1-4 In some embodiments, an alkyl group has 10-22 carbon atoms, also known as a C 10-22 In some embodiments, the alkyl group has 4-22 carbon atoms, also known as C 4-22 Alkylene.

[0235] The alkylene group may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the alkylene group is substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0236] The term "heteroalkylene" refers to the portion of a heteroalkane molecule that remains after removing two hydrogen atoms. In some embodiments, a heteroalkylene has 1-14 carbon atoms and is also referred to as a C 1-14 In some embodiments, a heteroalkylene group has 1-12 carbon atoms and is also referred to as a C 1-12 In some embodiments, the heteroalkylene group has 2-10 carbon atoms and is also referred to as a C 2-10 In some embodiments, heteroalkylene has 2-9 carbon atoms, also known as C 2-9 In other embodiments, the heteroalkylene group has one or more carbon atoms in the chain replaced by a heteroatom selected from S, O, and N. Exemplary heteroalkylene groups include etc.

[0237] Heteroalkylene may be unsubstituted or substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6In some embodiments, the heteroalkylene group is substituted by one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1- The alkyl group is substituted with a 6-membered alkoxy group or a 3- to 7-membered cyclo

[0238] Similarly, the definitions of "alkyleneoxy" and "alkenylene" are the same as those of "alkylene".

[0239] The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent, preferably a cycloalkyl ring containing 3 to 8 carbon atoms. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, and the like; polycyclic cycloalkyls include spirocyclic, paracyclic, and bridged cycloalkyls. Cycloalkyls may be unsubstituted or substituted with one or more radicals selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the cycloalkyl group is substituted with one or more halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0240] "Cycloalkenyl" refers to an unsaturated monocyclic or polycyclic hydrocarbon substituent containing one or more carbon-carbon double bonds. Preferably, the cycloalkenyl group contains 3 to 8 carbon atoms, such as a C3-C8 cycloalkenyl group. In addition, the cycloalkenyl group can be fused to an aryl or heteroaryl group. Exemplary cycloalkenyl groups include, but are not limited to:

[0241] Additionally, the cycloalkenyl group may be unsubstituted or substituted with one or more groups selected from halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 In some embodiments, the cycloalkenyl group is substituted with one or more halogen, hydroxy, oxo, cyano, amino, C 1-6 Alkyl, C 1-6 The alkyl group may be substituted with an alkoxy group or a 3- to 7-membered cycloalkyl group.

[0242] The terms "branched alkyl" and "branched alkenyl" refer to an alkyl or alkenyl group in which one carbon atom is bonded to at least two other carbon atoms, and not to form a cyclic group, such as a spirocycle. Examples: C6 alkyl groups include:

[0243] Similarly, the definitions of "alkyleneoxy", "alkenylene", "cycloalkylene" and "cycloalkenylene" are the same as those of "alkylene".

[0244] The term "hydroxy" refers to an -OH group.

[0245] The term "halogen" refers to fluorine, chlorine, bromine or iodine.

[0246] The term "amino" refers to -NH2.

[0247] The term "cyano" refers to -CN.

[0248] The term "oxo" refers to a =0 substituent.

[0249] "Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in a group are replaced independently of one another by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and a person skilled in the art can determine (by experiment or theory) whether substitution is possible or not without undue effort. BRIEF DESCRIPTION OF THE DRAWINGS

[0250] Figure 1: Luciferase mRNA lipid nanoparticles were injected into the tail vein of mice, and the fluorescence intensity of the mice was measured 24 hours later.

[0251] Figure 2: Luciferase mRNA lipid nanoparticles were injected into the tail vein of mice, and the fluorescence intensity of the mice was measured 48 hours later.

[0252] Figure 3: Fluorescence intensity of various organs and tissues of mice after tail vein injection of luciferase mRNA lipid nanoparticles.

[0253] Figure 4: Fluorescence intensity ratio of various organs and tissues of mice after tail vein injection of luciferase mRNA lipid nanoparticles.

[0254] Figure 5: Lipid nanoparticles containing luciferase mRNA were injected into the mice muscle, and the fluorescence intensity of the injected muscles was measured 24 hours later.

[0255] Figure 6: Mice were injected intramuscularly with hepatocyte growth factor mRNA lipid nanoparticles, and the protein expression level in the muscles of the injection site was detected 6 hours later.

[0256] Figure 7: After intramuscular injection of hepatocyte growth factor mRNA lipid nanoparticles into mice, the concentration of interleukin 6 (IL-6) in the injected muscle of mice was measured 6 hours later.

[0257] Figure 8: Protein expression in HEK293 cells after lipid nanoparticle delivery of mRNA.

[0258] Figure 9: Protein expression in HuH-7 cells after lipid nanoparticle delivery of mRNA.

[0259] Figure 10: Luciferase mRNA lipid nanoparticles were injected into the tail vein of mice, and the fluorescence intensity of the mice was measured 24 hours later.

[0260] Figure 11: Luciferase mRNA lipid nanoparticles were injected into the tail vein of mice, and the fluorescence intensity of the mice was measured 48 hours later. DETAILED DESCRIPTION

[0261] The present disclosure is further described below with reference to examples, but these examples are not intended to limit the scope of the present disclosure.

[0262] Experimental methods in the examples of this disclosure that do not specify specific conditions are generally based on conventional conditions or the conditions recommended by the raw material or product manufacturers. Reagents without specific sources are conventional reagents purchased from the market.

[0263] The structures of the compounds were determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). -6 The unit of ppm is given. NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), and deuterated methanol (Methanol-d4). The internal standard was tetramethylsilane (TMS).

[0264] HPLC determination used Agilent1100 high pressure liquid chromatograph, GAS15B DAD UV detector, Water Vbridge C18 150*4.6mm 5um chromatographic column.

[0265] MS was determined using an Agilent 6120 triple quadrupole mass spectrometer, a G1315D DAD detector, and a Waters Xbridge C18 4.6*50mm, 5um column. The samples were scanned in positive / negative ion mode with a mass scan range of 80-1200.

[0266] The thin layer chromatography silica gel plate used was Yantai Huanghai HSGF254 silica gel plate. The specification of the silica gel plate used in thin layer chromatography (TLC) was 0.2 mm ± 0.03 mm. The specification used for thin layer chromatography separation and purification products was 0.4 mm - 0.5 mm.

[0267] The flash column purification system used was Combiflash Rf150 (TELEDYNE ISCO) or Isolara one (Biotage).

[0268] Forward column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh or 300-400 mesh silica gel as the carrier, or uses Changzhou Santai pre-packed ultra-pure normal phase silica gel column (40-63μm, 60g, 24g, 40g, 120g or other specifications).

[0269] The known starting materials in the present disclosure can be synthesized by methods known in the art, or can be purchased from Shanghai Titan Technology, ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc, Bid Pharmaceuticals, etc.

[0270] Unless otherwise specified in the examples, all reactions were carried out under a nitrogen atmosphere.

[0271] Nitrogen atmosphere means that the reaction bottle is connected to a nitrogen balloon with a capacity of about 1L.

[0272] Hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.

[0273] Hydrogen was produced by a QPH-1L hydrogen generator from Shanghai Quanpu Scientific Instrument Co., Ltd.

[0274] The nitrogen atmosphere or hydrogen atmosphere is usually evacuated and filled with nitrogen or hydrogen, and the operation is repeated three times.

[0275] Unless otherwise specified in the examples, the solution refers to an aqueous solution.

[0276] Unless otherwise specified in the examples, the reaction temperature is room temperature, 20°C to 30°C.

[0277] The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent used in the reaction, the column chromatography eluent system used to purify the compound, and the developing solvent system for thin layer chromatography, the volume ratio of the solvent were adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid could be added for adjustment.

[0278] Example 1 Synthesis of Compound 1

[0279] Step 1:

[0280] Add water (46.8 mL) to a 500 mL three-necked flask. Cool to below 5°C in an ice-water bath, then slowly add sodium hydroxide (46.8 g, 1170 mmol). Once the sodium hydroxide is completely dissolved, add toluene (46.8 mL) and compound 1a (9.71 g, 50.0 mmol, purchased from Adamas). Stir at room temperature for 1 hour. Cool again to below 5°C in an ice-water bath, and slowly add compound 1b (34.1 g, 175 mmol, purchased from Adamas) dropwise. Heat to 25°C in an oil bath, stirring, and react until complete. The reaction mixture was cooled, water (200 mL) was added, and the mixture was extracted with methyl tert-butyl ether (100 mL × 3). The aqueous phase was cooled to below 5°C, concentrated hydrochloric acid was added until the pH reached about 3, and then extracted with DCM (100 mL × 5). The DCM phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography (EA / PE = 0-100%) to obtain 11.97 g of compound 1c (yield 94%).

[0281] 1 H NMR (400MHz, CDCl3): δ8.50(brs,1H),7.36-7.26(m,5H),4.51(s,2H),4.13~ 4.10(m,2H),3.56(t,2H),3.48(t,2H),1.67~1.61(m,4H),1.50~1.43(m,2H).

[0282] Step 2:

[0283] Compound 1c (21.7 g, 86.0 mmol) was weighed into a reaction flask and added sequentially with DCM (434 mL), compound 1d (19.9 g, 77.4 mmol, prepared according to a reference method, Tetrahedron, 2011, vol. 67, #2, pp. 303-311), DMAP (10.5 g, 30.1 mmol), and EDCI (19.8 g, 103 mmol). The mixture was allowed to react overnight at room temperature. TLC monitored the reaction completion. Water (500 mL) was added to the reaction mixture, and the layers were separated. The aqueous phase was extracted with DCM (300 mL). The combined organic phases were washed with saturated sodium bicarbonate solution and then with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography (EA / PE = 0-5%) to obtain 41.5 g of compound 1e (98% yield).

[0284] 1H NMR (400MHz, CDCl3): δ7.34~7.33(m,4H),7.28~7.27(m,1H),4.97~4.95(m,1H),4.50(s,2H),4.04 (s,2H),3.54~3.46(m,4H),1.69~1.62(m,4H),1.58~1.43(m,2H),1.25(s,26H),0.89~0.83(m,8H).

[0285] Step 3:

[0286] Compound 1e (41.5 g, 84.3 mmol) was weighed and dissolved in THF (208 mL). Pd(OH)2 / C (4.15 g, 20%) was added, and hydrogen was introduced through a double-layer hydrogen balloon. The reaction was stirred at room temperature. TLC monitored the reaction completion, and the mixture was filtered through celite. The filter cake was washed with EA (100 mL x 3), concentrated under reduced pressure, and purified by column chromatography (EA / PE = 50%) to obtain 27.0 g of compound 1f (80% yield).

[0287] 1 H NMR (400MHz, CDCl3): δ4.98~4.93(m,1H),4.05(s,2H),3.66(t,2H),3.54(t, 2H), 1.70~1.58(m,4H), 1.53~1.43(m,4H), 1.25(s,26H), 0.89~0.86(m,6H).

[0288] Step 4:

[0289] Compound 1f (6.01 g, 15.0 mmol) was weighed and dissolved in DCM (60 mL). Carbon tetrabromide (7.46 g, 22.5 mmol) was added under ice-cooling. After stirring for 10 minutes, triphenylphosphine (5.51 g, 21.0 mmol) was added and the reaction was stirred at room temperature. TLC monitoring indicated the reaction was complete. Purification by column chromatography (EA / PE = 0-5%) afforded 6.75 g of compound 1g (97% yield).

[0290] 1 H NMR (400MHz, CDCl3): δ4.99~4.93(m,1H),4.05(s,2H),3.54(t,2H),3.41(t,2H),1 .94~1.86(m,2H),1.70~1.63(m,2H),1.58~1.52(m,5H),1.26(s,25H),0.88(t,6H).

[0291] Step 5:

[0292] Compound 1g (1.0 g, 2.2 mmol) and compound 1h (1.9 g, 11.0 mmol, purchased from Anage) were dissolved in 20 mL of anhydrous ethanol, protected by nitrogen, heated to an internal temperature of 60°C for 16 hours, cooled to room temperature, and the reaction solution was concentrated under reduced pressure and purified by column chromatography (DCM:MeOH=10:1-5:1) to obtain 1.0 g of compound 1i (yield: 83%).

[0293] 1 H NMR (400MHz, CDCl3): δ5.13 (br s 1H),4.99~4.93(m,1H),4.04(s,2H),3.53(t,2H),3.25~3.16(m,2H),2.69(t,2H),2.63 (t,2H),1.71~1.61(m,4H),1.57~1.40(m,18H),1.33~1.19(m,24H),0.92~0.84(m,6H).

[0294] Step 6:

[0295] Compound 1i (0.487 g, 0.87 mmol), compound 1j (0.300 g, 0.795 mmol, prepared according to the method of patent application WO2018 / 170306), potassium iodide (0.145 g, 0.87 mmol), and potassium carbonate (0.438 g, 3.018 mmol) were added to a 25 mL flask, and 5 mL each of cyclopentane methyl ether and anhydrous acetonitrile were added. The system was protected by nitrogen and the heating reaction was basically complete. The reaction was cooled to room temperature, filtered, concentrated, and purified by column chromatography (PE:EA=10:1) to obtain 0.5 g of compound 1k (yield: 74%).

[0296] 1 H NMR (400MHz, CDCl3): δ5.60 (br s 1H),4.99~4.93(m,1H),4.84~4.78(m,1H),4.04(s,2H),3.53(t,2H),3.21 ~3.12(m,2H), 2.51~2.25(m,8H), 1.67~1.25(m,71H), 0.89~0.83(m,12H).

[0297] Step 7:

[0298] Compound 1k (0.5 g, 0.586 mmol) was dissolved in anhydrous dichloromethane (6.0 mL), and trifluoroacetic acid (2.0 mL) was added at room temperature for 4 hours. The mixture was concentrated under reduced pressure and purified by column chromatography (water:acetonitrile = 35%-95%) to obtain 370 mg of compound 1l (yield: 84%).

[0299] 1 H NMR (400MHz, CDCl3): δ4.99~4.93(m,1H),4.84~4.78(m,1H),4.04(s,2H),3.51(t,2H) ,2.72(t,2H),2.46~2.35(m,6H),2.29(t,2H),1.67~1.25(m,62H),0.89~0.83(m,12H).

[0300] Step 8:

[0301] Compound 1l (318 mg, 0.423 mmol) and compound 1m (59.6 mg, 0.423 mmol, prepared according to the literature method, Chemistry Letters, 2007, vol. 36, #8, p. 1012-1013) were added to a flask, and anhydrous ethanol (20 mL) was added. The system was protected by nitrogen and reacted at 25-35° C. for 18 hours. The mixture was concentrated under reduced pressure and purified by column chromatography (water: acetonitrile = 35%-100%) to obtain 290 mg of compound 1 (yield: 80%).

[0302] 1 H NMR (400MHz, CDCl3): δ4.96~4.92(m,1H),4.82~4.79(m,1H),4.05(s,2H),3.67(br s,2H),3.53(t,2H),3.27(d,3H),2.54(t,2H),2.46~2.40(m,4H),2.29(t,2H),1.79~1.29(m,62H),0.89~0.83(m,12H).

[0303] Example 2 Synthesis of Compound 2

[0304] Step 1:

[0305] Compound 1i (1.0 g, 1.8 mmol) and compound 2a (0.835 g, 1.8 mmol, prepared by the method of patent application WO2017 / 201333) were added to a flask, and potassium carbonate (0.996 g, 7.21 mmol) and potassium iodide (0.328 g, 1.98 mmol) were added. 10 mL each of anhydrous acetonitrile and cyclopentane methyl ether were added as solvents. The mixture was protected by nitrogen and heated until the reaction was basically complete. The mixture was cooled to room temperature, filtered, concentrated, and purified by column chromatography (PE:EA=10:1) to obtain 1.05 g of compound 2b (yield: 62%).

[0306] 1H NMR(400MHz, CDCl3): δ5.61(br s,1H),4.99~4.93(m,1H),4.89~4.83(m,1H),4.04(s,2H),3.52(t,2H),3.1 9~3.14(m,2H), 2.46~2.26(m,8H), 1.69~1.18(m,83H), 0.89~0.86(m,12H).

[0307] Step 2:

[0308] Compound 2b (1.05 g, 1.12 mmol) was added to a flask, and anhydrous dichloromethane (8 mL) was added under nitrogen protection. 4 M HCl / dioxane solution (4 mL) was added at room temperature and reacted for 4 hours. The crude product was concentrated under reduced pressure and dissolved in 50 mL of dichloromethane solution. The product was washed twice with saturated aqueous sodium carbonate solution and once with 20 mL of 0.1 N NaOH aqueous solution. The product was dried over anhydrous sodium sulfate, filtered, concentrated, and purified by column chromatography (acetonitrile:methanol = 0-100%) to obtain 0.85 g of compound 2c (yield: 91%).

[0309] 1 H NMR (400MHz, CDCl3): δ4.99~4.93(m,1H),4.89~4.83(m,1H),4.04(s,2H),3.52(t,2H) ,2.72(t,2H),2.46~2.35(m,6H),2.27(t,2H),1.69~1.18(m,74H),0.89~0.82(m,12H).

[0310] Step 3:

[0311] Compound 2c (0.25 g, 0.298 mmol) was added to a flask, followed by compound 1m (0.042 g, 0.298 mmol), and anhydrous ethanol (5 mL). The mixture was reacted at room temperature for 16 hours, concentrated under reduced pressure, and purified by column chromatography (acetonitrile:methanol = 0-100%) to obtain 0.25 g of compound 2 (yield: 89%).

[0312] 1H NMR (400MHz, CDCl3): δ8.69(br s,1H),8.22(br s,1H),4.96~4.90(m,1H),4.89~4.82(m,1H),4.03(s,2H),3.76~3.67(m,2H),3.53(t,2H),3.31(s,3H),3.2 8~3.20(m,2H),3.16~2.99(m,4H),2.28(t,2H),2.15~2.04(m,2H),1.83~1.17(m,72H),0.91~0.82(m,12H).

[0313] Example 3 Synthesis of Compound 3

[0314] Compound 3b (0.4 g, 0.532 mmol, prepared by the method of patent application WO2018 / 170306) and compound 3a (37.8 mg, 0.266 mmol, prepared by the literature method, Organic Syntheses, 1999, vol. 76, p. 189-189) were added to a flask, and anhydrous ethanol (18 mL) was added. The mixture was protected by nitrogen and reacted at room temperature for 16 hours. The mixture was concentrated under reduced pressure and purified by column chromatography (acetonitrile: methanol = 0-100%) to obtain 0.19 g of compound 3 (yield: 45%).

[0315] 1 H NMR (400MHz, CDCl3): δ7.11(br s,2H),4.89~4.77(m,4H),3.73~3.58(m,4H),2.61~2.52(m,4H),2.46~2 .36(m,8H),2.32~2.24(m,8H),1.78~1.17(m,132H),0.93~0.79(m,24H).

[0316] Example 4 Synthesis of Compound 4

[0317] Step 1:

[0318] Compound 4a (2.3 g, 10.1 mmol, purchased from Leyan) and compound 2a (1.0 g, 2.1 mmol) were added to a flask, and anhydrous ethanol (60 mL) was added under nitrogen protection. The reaction was heated for 18 hours, and then the heating was stopped. The mixture was cooled to room temperature, concentrated, and purified by column chromatography (DCM:MeOH=10:1) to obtain 0.85 g of compound 4b (yield: 65%).

[0319] 1H NMR (400MHz, CDCl3): δ4.89~4.83(m,1H), 4.71~4.33(m,1H), 3.71~3.38(m,1H), 2.62~2. 41(m,3H),2.29~2.25(m,2H),2.03~1.93(m,2H),1.70~1.10(m,54H),0.89~0.84(m,6H).

[0320] Step 2:

[0321] Compound 4b (400 mg, 0.67 mmol) and compound 1j (254 mg, 0.67 mmol) were added to a flask, followed by potassium carbonate (371.5 mg, 2.69 mmol) and potassium iodide (123.0 mg, 0.74 mmol), and 5 mL each of anhydrous acetonitrile and cyclopentane methyl ether. The mixture was protected by nitrogen and heated until the reaction was almost complete. The mixture was cooled to room temperature, filtered, concentrated, and purified by column chromatography (PE:EA=10:1) to obtain 211 mg of compound 4c (yield: 35%).

[0322] MS m / z(ESI):891.7[M+H] + .

[0323] Step 3:

[0324] Compound 4c (0.211 g, 0.237 mmol) was dissolved in anhydrous dichloromethane (1.5 mL), and trifluoroacetic acid (1.5 mL) was added. The mixture was reacted at room temperature for 4 hours, concentrated under reduced pressure, and purified by column chromatography (water:acetonitrile = 35%-95%) to obtain 126 mg of compound 4d (yield: 67%).

[0325] 1 H NMR (400MHz, CDCl3): δ4.89~4.78(m,2H), 2.47~2.34(m,4H), 2.31~2.24(m,4H), 1.68~1.18(m,76H), 0.92~0.83(m,12H).

[0326] Step 4:

[0327] Compound 4d (81 mg, 0.102 mmol) and compound 1m (29.2 mg, 0.206 mmol) were added to a flask, and anhydrous ethanol (5 mL) was added. The system was protected by nitrogen and reacted at 25-35°C. TLC showed that the reaction was complete. The mixture was concentrated under reduced pressure and purified by column chromatography (water:acetonitrile = 35%-95%) to obtain 88 mg of compound 4 (yield: 95%).

[0328] 1H NMR (400MHz, CDCl3): δ4.89~4.77(m,2H), 2.37~2.28(m,3H), 2.56~2.24(m,8H), 2.15~2.06(m,1H), 1.84~1.18(m,75H), 0.93~0.81(m,12H).

[0329] Example 5

[0330] Compound 5:

[0331] MS (ESI): m / z 862.68 [M+H] + .

[0332] 1 H NMR (400MHz, CDCl3): δ4.78~4.85(m,2H), 3.66~3.68(m,2H), 3.42~3.47(m,4H), 3.27~3.29(m,3H) ,2.49~2.54(m,4H),2.36~2.40(m,4H),2.27~2.31(m,2H),1.20~1.80(m,62H),0.85~0.89(m,12H).

[0333] Compound 6:

[0334] MS (ESI): m / z 862.68 [M+H] + .

[0335] 1 H NMR (400MHz, CDCl3): δ4.79~4.86(m,2H), 3.28~3.58(m,9H), 2.60~2.62(m,4H) ,2.42~2.46(m,2H),2.28~2.34(m,4H),1.20~1.80(m,62H),0.86~0.89(m,12H).

[0336] Comparative Example 1

[0337] Prepared according to the method in US10392341

[0338] Comparative Example 2

[0339] Prepared according to the method in WO2023125738

[0340] Comparative Example 3

[0341] Prepared according to the method in US9868692

[0342] Example 6: Preparation of lipid nanoparticle composition

[0343] 1.1 Preparation method

[0344] Compounds 1-6 and 1-3 were dissolved in ethanol and mixed with ethanolic DSPC, cholesterol, and DMG-PEG solutions at a molar ratio of 50:10:38.5:1.5 to prepare ethanolic lipid solutions. Luciferase mRNA (GenBank accession number: MN728548.1) was dissolved in 50 mM acetate buffer, pH 5, to prepare an mRNA aqueous solution (0.2 mg / mL). The ethanolic lipid solution and mRNA aqueous solution were mixed using microfluidics at a volume ratio of 1:3 for the ethanol phase and a total lipid to mRNA weight ratio of approximately 20:1 to prepare lipid nanoparticles. The solution was dialyzed against 20 mM Tris, pH 7.5, to remove ethanol, and finally exchanged into 20 mM Tris, pH 7.5, 8% sucrose solution for cryopreservation to obtain mRNA-encapsulated lipid nanoparticle compositions.

[0345] 1.2 Characterization of lipid nanoparticle composition

[0346] The nanoparticle size and polydispersity index (PDI) were determined by dynamic light scattering using a Malvern Zetasizer Pro in 173° backscattering detection mode.

[0347] The liposome encapsulation efficiency was determined using the Quant-iT RiboGreen RNA Assay Kit.

[0348] Table 1

[0349] Test Example 1: Evaluation of mRNA delivery efficiency and tissue distribution of lipid nanoparticle compositions via tail vein injection in vivo

[0350] Female Balb / c mice aged 6-8 weeks were injected intravenously with a lipid nanoparticle composition encapsulating luciferase mRNA (GenBank Accession No.: MN728548.1) at a dose of 0.5 mg / kg. Luciferase substrate was injected intraperitoneally into each mouse 24 and 48 hours after administration. Fluorescence images of the tissues were taken using an IVIS Small Animal Optical Live Imager (PerkinElme), and the fluorescence intensity was calculated. Fluorescence intensity represents the expression of luciferase protein, reflecting the efficiency of mRNA delivery by the lipid nanoparticles in vivo. After 24 hours, the mice were dissected and the heart, liver, spleen, lung, and kidney were obtained. Images were taken using an IVIS Small Animal Optical Live Imager, and the fluorescence intensity of each tissue was calculated. The mean fluorescence intensity was calculated for at least three biological replicates of each lipid nanoparticle composition for each compound. The data are shown in Table 2, Figures 1, and 2. The fluorescence intensities in Figures 1 and 2 represent the fluorescence intensities of each mouse tissue as measured by the IVIS Small Animal Optical Live Imager. Fluorescence intensity represents the level of luciferase protein expression, indicating the efficiency of mRNA delivery and tissue distribution of the lipid nanoparticle composition in vivo. * in Figures 1 and 2 indicates statistically significant differences between groups at P < 0.05.

[0351] The lipid nanoparticle compositions corresponding to comparative compounds 1 and 2 were used as controls to detect the mRNA delivery efficiency of liposome nanoparticle compositions corresponding to compounds 1, 2, 3 and 4 injected into the tail vein of mice.

[0352] Table 2

[0353] Conclusion: As shown in Table 2, Figures 1 and 2, the fluorescence intensity of the lipid nanoparticle compositions composed of compounds 1-4 was significantly higher than that of the lipid nanoparticle compositions corresponding to comparative compounds 1 and 2 at 24 and 48 hours in mice. This indicates that the lipid nanoparticles corresponding to compounds 1 to 4 and their compositions have better delivery efficiency in vivo.

[0354] As shown in Table 3 and Figure 4, the fluorescence intensity and proportion represent the protein expression amount in various organ tissues and the proportion in each tissue. The lipid nanoparticle composition composed of compound 2 has the highest fluorescence intensity in the liver, accounting for more than 95%, indicating that the lipid nanoparticle composition corresponding to this type of compound has good liver targeting ability.

[0355] Table 3

[0356] Test Example 2: Evaluation of the mRNA delivery efficiency of lipid nanoparticle compositions by intramuscular administration

[0357] Female Balb / C mice aged 6-8 weeks were intramuscularly administered with lipid nanoparticles encapsulating luciferase mRNA (GenBank accession number: MN728548.1) at a dose of 0.1 mg / kg. 24 hours after administration, each mouse was intraperitoneally injected with luciferase substrate, and fluorescence images were taken using an IVIS small animal optical in vivo imaging instrument (PerkinElme) to measure the fluorescence intensity of each tissue. The fluorescence intensity represents the expression level of luciferase protein. The average fluorescence intensity was calculated for at least 3 biological replicates of the lipid nanoparticle composition corresponding to each compound. The data are shown in Table 4 and Figure 5. * in Figure 5 indicates statistical P<0.05, indicating that there is a statistically significant difference between the groups.

[0358] The lipid nanoparticle compositions corresponding to comparative compound 1 and comparative compound 2 were used as controls to detect the mRNA delivery efficiency of compound 1, compound 2, and the corresponding lipid nanoparticle compositions in mice by intramuscular injection.

[0359] Conclusion: As shown in Table 4 and Figure 5, the fluorescence intensity of the lipid nanoparticle compositions corresponding to Compound 1 and Compound 2 in mice was significantly higher than that of the lipid nanoparticle compositions corresponding to Comparative Compound 1 and Comparative Compound 2. This indicates that the lipid nanoparticles corresponding to Compound 1 and Compound 2 and their compositions have better delivery efficiency.

[0360] Table 4

[0361] A 0.05 mg / kg dose of lipid nanoparticles containing mRNA expressing hepatocyte growth factor (HGF) (NCBI Accession No. NM_000601.6) was injected into the gastrocnemius muscle of 6-8 week-old female BALB / c mice. Six hours after injection, the muscle was harvested, ground, and lysed. HGF protein expression (ng / mg) was measured using an ELISA kit. This assay measures the amount of HGF protein expressed per unit of total muscle tissue protein. The average protein concentration was calculated for each compound using at least three replicates of the lipid nanoparticle composition.

[0362] The mRNA delivery efficiency of lipid nanoparticle compositions corresponding to Compounds 1 and 2 was tested by intramuscular injection, using the lipid nanoparticle composition corresponding to Compound 3 as a control. The relevant data are shown in Figure 6 and Table 5.

[0363] In Figure 6, * indicates statistical P<0.05, indicating that there are statistically significant differences between the groups.

[0364] Table 5

[0365] Conclusion: As shown in Table 5 and Figure 6, the protein expression levels of Compounds 1 and 2 were significantly higher than those of the comparative compound 3, indicating that the lipid nanoparticle compositions corresponding to Compounds 1 and 2 were significantly more effective than the comparative compound 3 in delivering mRNA expressing hepatocyte growth factor via intramuscular injection in vivo. Compound 1 was also significantly better than Compound 2, indicating that the lipid nanoparticle compositions corresponding to Compounds 1 and 2 were more efficient in delivering mRNA and expressing protein in muscle.

[0366] Test Example 3: Evaluation of the in vivo safety of lipid nanoparticle compositions

[0367] Foreign substances entering the mammalian body trigger an innate immune response, thereby promoting the production of cytokines. These substances, upon entry, trigger an inflammatory response, which can easily lead to adverse reactions such as fever and edema. Therefore, the immunogenicity of lipid nanoparticle compositions in vivo was assessed by evaluating the concentration of cytokines, such as interleukin-6 (IL-6), in the blood of mice after injection with the lipid nanoparticle compositions. Lower cytokine concentrations indicate that the lipid nanoparticle compositions have lower immunogenicity and, therefore, better biosafety.

[0368] 6-8 week old female BALB / c mice were injected intramuscularly with a lipid nanoparticle composition encapsulating mRNA expressing hepatocyte growth factor at a dose of 0.05 mg / kg. Six hours after injection, muscle tissue homogenates were collected. Muscle tissue IL-6 concentrations were measured using a mouse IL-6 ELISA kit (pg / mg of tissue protein). Serum IL-6 concentrations were measured in at least three biological replicates for each lipid nanoparticle composition, and the mean value was calculated.

[0369] Conclusion: As shown in Table 6 and Figure 7, the lipid nanoparticle composition corresponding to Compound 1 significantly lowered IL6 concentrations in mouse muscle tissue than the control compounds 1 and 2. This suggests that the lipid nanoparticle composition containing Compound 1 has lower immunogenicity and better biosafety in vivo. ** in Figure 7 indicates statistically significant differences, P < 0.01.

[0370] Table 6

[0371] Test Example 4: Evaluation of Lyophilization of Lipid Particle Compositions

[0372] Lipid nanoparticles were prepared according to the method in Example 1 and ultrafiltration was performed into a buffer solution containing 20 mM Tris, pH 7.5, and 8% sucrose to obtain a lyophilized solution of the lipid nanoparticle composition. The lyophilized solution of the lipid nanoparticle composition was then filled into 2 mL vials at a rate of 0.6 mL per vial, with an mRNA concentration of 0.2 mg / mL per vial. The solution was then placed in a lyophilizer. Pre-freezing at -45°C for 3 hours, followed by a primary drying at -40°C for 30 hours, and a secondary drying at 30°C for 10 hours, all under a vacuum of 0.05 mmbar, was completed. The final lyophilized product was obtained upon removal from the freezer.

[0373] The lyophilized product was reconstituted with 0.6 mL of water for injection and the lipid nanoparticle size and encapsulation efficiency were measured using the aforementioned methods. The particle sizes of the lipid nanoparticles before and after lyophilization were compared. The data are as follows:

[0374] Table 7

[0375] The particle size of the lipid nanoparticles corresponding to compound 1 before and after lyophilization is significantly lower than that of the comparative compound 2 and compounds 5 and 6. The particle size growth after lyophilization and reconstitution is significantly smaller than that of compound 5, compound 6 and comparative compound 2. The particle size distribution (PDI) of the lipid nanoparticles corresponding to compound 1 is less than 0.1, which is significantly lower than that of compounds 5 and 6 and comparative compound 2. This shows that the particle size distribution of the lipid nanoparticles corresponding to compound 1 is more uniform and can still maintain a uniform particle size distribution after lyophilization. The encapsulation efficiency of compound 1 after reconstitution after lyophilization is still higher than 90%, while the encapsulation efficiency of the lipid nanoparticles corresponding to comparative compounds 2, compounds 5 and 6 after reconstitution after lyophilization is significantly reduced, which is lower than the general encapsulation efficiency quality control standard of 85%.

[0376] This indicates that the lipid nanoparticles corresponding to compound 1 and their compositions can withstand freeze-thaw and temperature changes during the freeze-drying process and are suitable for the freeze-drying process.

[0377] Test Example 5: Evaluation of mRNA delivery efficiency of lipid nanoparticle compositions on cells

[0378] Detection of protein expression of liposome nanoparticle compositions corresponding to compounds 1, 5 and 6 after delivery of mRNA in cells

[0379] HEK 293 cells were seeded into 96-well plates and cultured overnight. When the cell density reached more than 80%, a lipid nanoparticle composition solution encapsulating luciferase mRNA (GenBank accession number: MN728548.1) was added to the cell plate well culture medium at an mRNA dose of 100 ng / well. After 24 hours, the fluorescence intensity of the expressed luciferase protein was detected using a luciferase reporter gene assay kit (Promega) and a microplate reader. The fluorescence intensity value, i.e., the fluorescence value detected by the microplate reader, represents the expression level of the luciferase protein. The higher the intensity of the strong light, the higher the protein expression level. The average fluorescence intensity was calculated for at least 3 groups of lipid nanoparticles corresponding to each compound.

[0380] Table 8

[0381] Conclusion: As shown in the table and Figure 8, the expression level of the lipid nanoparticle composition composed of compound 1 was significantly higher than that of compound 5 and compound 6, indicating that compound 1 has better delivery efficiency.

[0382] In Figure 8, *** represents statistically significant differences, P < 0.001.

[0383] HuH-7 cells (human liver cancer cells) were seeded into 96-well plates and cultured overnight. When the cell density reached more than 80%, a lipid nanoparticle composition solution encapsulating luciferase mRNA (GenBank accession number: MN728548.1) was added to the cell plate well culture medium at an mRNA dose of 100 ng / well. After 24 hours, the fluorescence intensity of the expressed luciferase protein was detected using a luciferase reporter gene assay kit (Promega) and a microplate reader. The fluorescence intensity value, i.e., the fluorescence value detected by the microplate reader, represents the expression level of the luciferase protein. The higher the intensity of the strong light, the higher the protein expression level. The average fluorescence intensity was calculated for at least 3 groups of lipid nanoparticle compositions corresponding to each compound.

[0384] Table 9

[0385] Conclusion: As shown in Table 9 and Figure 9, the expression level of the lipid nanoparticle composition composed of Compound 1 was significantly higher than that of Compound 5, Compound 6, and Comparative Compound 1, indicating that Compound 1 has better delivery efficiency. The expression level of the lipid nanoparticle composition composed of Compound 5 was not significantly different from that of Comparative Compound 1.

[0386] In FIG9 , ** indicates statistical P<0.01, *** indicates statistical P<0.001, and indicates that there are statistically significant differences between the groups.

[0387] Test Example 6: Evaluation of the mRNA delivery efficiency of lipid nanoparticle compositions via tail vein injection in vivo

[0388] Female Balb / c mice aged 6-8 weeks were injected with a lipid nanoparticle composition encapsulating luciferase-expressing mRNA (GenBank Accession No.: MN728548.1) via the tail vein at a dose of 0.5 mg / kg. Luciferase substrate was injected intraperitoneally into each mouse 24 and 48 hours after administration. Fluorescence images were taken of the tissues using an IVIS small animal optical in vivo imaging instrument (PerkinElme), and the fluorescence intensity was calculated. Fluorescence intensity represents the expression of luciferase protein, which in turn reflects the efficiency of mRNA delivery by the lipid nanoparticles in vivo. The mean fluorescence intensity was calculated for at least three biological replicates of each lipid nanoparticle composition for each compound. The data are shown in Table 10, Figures 10, and 11. The fluorescence intensity in Figures 10 and 11 represents the fluorescence intensity corresponding to each mouse tissue as captured and calculated using the IVIS small animal optical in vivo imaging instrument. Fluorescence intensity represents the expression of luciferase protein, which in turn indicates the efficiency and tissue distribution of mRNA delivery by the lipid nanoparticle composition in vivo.

[0389] In Figures 10 and 11, * indicates that P<0.05, indicating that there is a statistically significant difference between the groups. ns indicates that P≥0.05, indicating that there is no statistically significant difference between the groups.

[0390] The mRNA delivery efficiency of the liposome nanoparticle compositions corresponding to Compound 5 and Compound 6 was tested by tail vein injection in mice, using the lipid nanoparticle composition corresponding to Compound 1 as a control.

[0391] Table 10

[0392] Conclusion: As shown in Table 10, Figure 10 and Figure 11, the fluorescence intensity of the lipid nanoparticle composition composed of compound 5 in mice was significantly lower than that of the lipid nanoparticle composition corresponding to the control compound 1 at 24 hours and 48 hours.

Claims

1. A compound represented by formula I or a salt thereof, in, H 1 Selected from C 1-12 Alkylene, C 2-12 Alkenylene, C 3-8 Cycloalkylene or C 3-8 Cycloalkenylene; H 2 Selected from C 1-12 Alkylene or C 1-12 heteroalkylene; The condition is that when H 2 C 1-12 When alkylene is present, H 1 Selected from C 3-8 Cycloalkylene or C 3-8 Cycloalkenylene; R 1 Selected from C 1-24 Alkyl or C 2-24 alkenyl; R 2 Selected from C 1-24 Alkyl, C 1-24 Alkenyl or -R b -L 2 -R c ; L 1 or L 2 Each is independently selected from -C(O)O-, -OC(O)-, -C(O)-, -OC(O)O-, -O-, -S(O)q-, -SS-, -C(O)S-, -SC(O)-, -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a -、-P(O)(OR a )-or key, R a Selected from hydrogen or C 1-6 Alkyl, q is selected from 0, 1 or 2; R b Selected from C 1-12 Alkylene or C 2-12 alkenylene; R c Selected from C 1-24 Alkyl or C 2-24 alkenyl; R 3 Selected from alkylamino or cycloalkylamino; R 4 Selected from hydrogen or C 1-6 alkyl.

2. The compound or salt thereof according to claim 1, wherein H 2 C 1-12 Heteroalkylene, preferably C 2- 9 heteroalkylene.

3. The compound or salt thereof according to claim 1 or 2, wherein R 3 It is an alkylamino group, preferably a methylamino group or a dimethylamino group.

4. The compound or salt thereof according to any one of claims 1 to 3, wherein R 1 C 2-24 Alkyl, preferably C 4-18 Alkyl; or R 1 C 2-24 Alkenyl, preferably C 4-18 Alkenyl.

5. The compound or salt thereof according to any one of claims 1 to 4, wherein R 2 For-R b -L 2 -R c , R b , L 2 and R c As defined in claim 1.

6. The compound or salt thereof according to claim 1 or 5, wherein the compound represented by formula I is a compound represented by formula IIa Where X is -N(R d )- or -O-, o is selected from an integer between 1 and 11, p is selected from an integer between 1 and 5, and o+p is not greater than 12, R d Selected from hydrogen or C 1-6 Alkyl, R 5 , R 6 , R 7 , R 8 are each independently selected from hydrogen, halogen, hydroxyl or C 1-6 Alkyl, L 1 , L 2 , H 1 , R 1 , R 3 , R 4 , R b and R c As defined in claim 1.

7. The compound or salt thereof according to claim 6, wherein p is selected from 1 or 2.

8. The compound or salt thereof according to claim 6, wherein o is selected from an integer between 4 and 7, such as 5 or 6.

9. The compound or salt thereof according to any one of claims 1 to 8, wherein H 1 C 1-12 Alkylene, preferably C 3-8 Alkylene.

10. The compound or salt thereof according to claim 1, wherein H 1 Selected from C 3-8 Cycloalkylene, preferably the compound represented by formula I is a compound represented by formula IIb Where L 1 , L 2 , H 2 , R 1 , R 3 , R 4 , R b and R c As defined in claim 1.

11. The compound or salt thereof according to any one of claims 1 to 10, wherein L 1 or L 2 are each independently selected from -C(O)O-, -OC(O)- or a bond.

12. The compound or salt thereof according to claim 1, 6 or 10, wherein R b C 1-12 Alkylene.

13. The compound or salt thereof according to claim 1, 6 or 10, wherein R c C 1-24 alkyl.

14. The compound represented by formula A or its salt Among them, M 1 Each independently selected from C 1-12 Alkylene, C 2-12 Alkenylene, C 3-8 Cycloalkylene or C 3-8 Cycloalkenylene; M 2 Each independently selected from C 1-12 Alkylene or C 1-12 heteroalkylene; R 1 Each independently selected from C 1-24 Alkyl or C 2-24 alkenyl; R 2 Each independently selected from C 1-24 Alkyl, C 1-24 Alkenyl or -R b -L 2 -R c ; L 1 or L 2 Each is independently selected from -C(O)O-, -OC(O)-, -C(O)-, -OC(O)O-, -O-, -S(O)q-, -SS-, -C(O)S-, -SC(O)-, -NR a C(O)-、-C(O)NR a -、-NR a C(O)NR a -、-NR a C(O)O-、-OC(O)NR a -、-P(O)(OR a ) or key, R a Selected from hydrogen or C 1-6 Alkyl or C 2-6 alkenyl, q is selected from 0, 1 or 2; R b Each independently selected from C 1-12 Alkylene or C 2-12 alkenylene; R c Each independently selected from C 1-24 Alkyl or C 2-24 alkenyl; R 4 are each independently selected from hydrogen or C 1-6 alkyl.

15. The compound or salt thereof according to any one of claims 1 to 14, wherein R 1 or R c Selected from:

16. The compound or salt thereof according to claim 1 or claim 14, wherein the compound represented by formula I is selected from:

17. An isotope substitution product of the compound according to any one of claims 1 to 16, preferably, the isotope substitution product is a deuterium atom substitution product.

18. A lipid particle comprising the compound according to any one of claims 1 to 16 or a salt thereof or an isotope substitute thereof, and further comprising an active agent, wherein the active agent is preferably a polynucleotide or a nucleic acid, such as DNA, siRNA or mRNA.

19. A pharmaceutical composition comprising the lipid particles according to claim 18 and a pharmaceutically acceptable excipient.

20. Use of a compound or salt thereof according to any one of claims 1 to 16, or an isotope substitute according to claim 17, or a lipid particle according to claim 18, or a pharmaceutical composition according to claim 19 in the preparation of a medicament for preventing and / or treating cancer, infection, autoimmune disease, neurodegenerative disease and inflammation.

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