WO2024103225A1 - Cationic polyester, preparation method therefor and use thereof - Google Patents

Abstract

A cationic polyester, a preparation method therefor and the use thereof. The cationic polyester is a hyperbranched polymer, which is formed by polymerizing three monomers, monomer P, monomer S and monomer M, or two monomers, the monomer P and monomer S, with monomer T, an end group of said hyperbranched polymer being modified with group E. The monomer P is a lactone. The monomer S is an organic acid of which an end group contains two or more carboxyl groups. The monomer M is a compound containing two hydroxyl groups and one secondary amino group or tertiary amino group. The monomer T is a compound containing three or more hydroxyl groups. An end group modifying compound E which provides the group E modification is a compound containing at least one primary amino group, secondary amino group or tertiary amino group. When being used for nucleic acid drug delivery, the cationic polyester can remarkably improve the nucleic acid transfection efficiency and has lower cytotoxicity, and thus can be used clinically. In addition, the cationic polyester has low preparation cost and is environment-friendly and pollution-free.

Description

Cationic polyester and its preparation method and application Technical Field

The present application relates to the technical field of nucleic acid delivery materials, and in particular to a cationic polyester and a preparation method and application thereof.

Background technique

Gene therapy based on nucleic acid drug delivery is believed to lead the third industrial revolution in biomedicine. Among them, compared with other technologies such as DNA or viral vectors, mRNA technology has the advantages of high efficiency, high safety, shorter production cycle and lower production cost. mRNA technology has become famous in the field of new crown vaccines, and it is expected to bring new methods to many fields such as cancer and immune diseases. However, due to the instability of mRNA molecules and low in vivo delivery efficiency, the application of this technology has been limited. To achieve the widespread application of mRNA technology, it is necessary to have a suitable delivery carrier to deliver it to the body in order to have a practical effect. At present, the commercially successful delivery technology is mainly the lipid nanoparticle (LNP) technology based on ionizable cationic lipids used by companies such as Moderna and Biotech. Although LNP has been widely used in new crown vaccines; however, the relatively low efficiency and significant side effects of LNP greatly limit its scope of application. Therefore, how to develop a new, efficient and non-toxic delivery system is still a bottleneck problem that inhibits the development of mRNA technology.

Yale University in the United States previously reported a biodegradable cationic polyester that can be used for DNA and mRNA delivery and its preparation method. The preparation method is based on in vitro synthetic biology technology. The linear polymer is obtained by catalyzing the polymerization of three monomers by immobilized lipase. It is used directly or end-group modified to obtain a biodegradable cationic polyester that can be used for DNA and mRNA delivery. Compared with commercial reagents, Yale University's cationic polyester has higher transfection efficiency and lower toxicity. However, the transfection efficiency of the cationic polyester is still insufficient, and it needs to be dissolved in toxic organic solvents, making it difficult to use in actual clinical practice.

Therefore, how to develop nucleic acid delivery materials with higher transfection efficiency and non-toxicity or lower toxicity remains the research focus and difficulty in this field.

Summary of the invention

The purpose of this application is to provide a new cationic polyester and its preparation method and application.

In order to achieve the above objectives, this application adopts the following technical solutions:

The first aspect of the present application discloses a cationic polyester, which is a hyperbranched polymer formed by polymerizing three monomers, monomer P, monomer S and monomer M, or two monomers, monomer P and monomer S, with monomer T, and the end group of the hyperbranched polymer is modified with a group E; the monomer P is a cyclic lactone; the monomer S is an organic acid having two or more carboxyl groups at the end group; the monomer M is a compound containing two hydroxyl groups and one secondary amine or tertiary amine; the monomer T is a compound containing three or more hydroxyl groups; the end group modified compound E provided with the group E modification is a compound containing at least one primary amine, secondary amine or tertiary amine group.

It should be noted that the polymerization of the three monomers P, S, and M to obtain a linear polymer and the modification of the end group E is a technology that has been reported by Yale University; based on this, the present application has found that by adding a monomer T to the three monomers, or directly replacing the monomer M with monomer T, a new polymer with a hyperbranched structure can be obtained by polymerization, i.e., the new cationic polyester of the present application. In addition, the study found that when the cationic polyester of the present application is used for nucleic acid delivery, it can not only significantly improve the gene transfection efficiency, but also has lower cytotoxicity, which can better meet clinical use.

In one implementation of the present application, the monomer P is a cyclic lactone with a fatty chain length of 6 to 35.

In one implementation of the present application, the monomer P is selected from but not limited to at least one of cyclohexyl lactone, cyclododecanolide, cyclopentadecanolide and cyclohexadecanolide.

In one implementation of the present application, the monomer S is an organic acid with a carbon chain length of 3 to 18.

In one implementation of the present application, the monomer S is selected from but not limited to at least one of adipic acid, sebacic acid and 1,2,3-propanetricarboxylic acid.

In one implementation of the present application, the monomer M is a compound having a carbon chain length of 4 to 36 and containing two hydroxyl groups and a secondary amine or a tertiary amine.

In one implementation of the present application, the monomer M is selected from but not limited to at least one of diethanolamine, methyldiethanolamine and ethyldiethanolamine.

In one implementation of the present application, monomer T is a compound having a carbon chain length of 4 to 54 and containing three or more hydroxyl groups.

In one implementation of the present application, monomer T is selected from but not limited to at least one of trimethylolpropane, 3-(hydroxymethyl)-1,5-pentanediol, triethanolamine, and N,N,N’,N’-tetrahydroxyethylethylenediamine.

In one implementation of the present application, the terminal group-modified compound E modified with the group E is at least one of E1 to E26.

Preferably, the terminal group modified compound E is at least one of E1 to E10, E12, E14 to E21, E25, and E26.

More preferably, the terminal group modified compound E is at least one of E2, E4, E9, E10, E12, E14, E15, E25, and E26.

It should be noted that the cationic polyesters modified with end groups using end group-modified compounds E1 to E26 in the present application all have good transfection efficiency and low cytotoxicity; in particular, the transfection efficiency of cationic polyesters modified with end groups of E2, E4, E9, E10, E12, E14, E15, E25, and E26 is significantly better than that of other end group-modified cationic polyesters, and is also significantly better than the currently most efficient commercial transfection reagent LipoMM.

In one implementation of the present application, the cationic polyester is a hyperbranched polymer having a structure shown in Formula 1;

Formula 1

Wherein, x, y, and z are independent integers ranging from 1 to 200.

n is an integer from 0 to 200,

j and k are integers from 0 to 30,

l, m, o, p, q are independent integers from 1 to 20,

_Rx is hydrogen, or a substituted or unsubstituted alkyl group containing 1 to 18 carbon atoms, or a substituted or unsubstituted aromatic group containing at least one benzene ring, or a substituted or unsubstituted heterocyclic group containing at least one heterocyclic ring, or a substituted or unsubstituted alkoxy group containing 1 to 18 carbon atoms and at least one oxygen atom;

J is hydrogen, R ₁ is modified by the group E provided by the terminal group modification compound E, and there is no R ₂ ; or, J is a carbonyl group, and both R ₁ and R ₂ are modified by the group E provided by the terminal group modification compound E;

In Formula 1, the truncation line represents a branch structure, the first monomer connected to the branch structure is P or S, and other monomers are subsequently connected to form a hyperbranched structure.

Herein, connecting other monomers means, for example, subsequently connecting monomers in the order of formula 1 to form a branched structure based on the branched structure, and so on, to form a dendrite-like hyperbranched structure.

It should be noted that _R1 and/or _R2 in the present application are group E modifications provided by terminal group modification compound E. When terminal group modification compound E performs terminal group E modification on hyperbranched polymer, terminal group modification compound E will reduce one hydrogen to form a terminal group of group E combined with hyperbranched polymer. When N,N'-carbonyldiimidazole (CDI) is used as a coupling agent, terminal group modification of group E will be formed at both ends of the hyperbranched polymer shown in formula 1 at the same time; at this time, in order to connect group E to the hyperbranched polymer, CDI will provide a carbonyl group at the position of terminal group _R2 for connecting group E and hyperbranched polymer; at the position of _R1 , terminal group modification compound E will still reduce one hydrogen to form group E, which is directly connected to the hyperbranched polymer.

It should also be noted that when only monomers P and S are polymerized with monomer T to form a hyperbranched polymer, that is, n is 0, if J is a carbonyl group, one end of the carbonyl group is connected to R ₂ , and the other end is connected to the hydroxyl group of monomer T to form a terminal group E modification. It can be understood that in this case, the structure of the general formula described in Formula 1 will be adaptively adjusted, that is, monomer T is connected to the carbonyl group J at the end.

In one implementation of the present application, the number average molecular weight of the cationic polyester is 1-30k, preferably 2-20k.

The second aspect of the present application discloses a method for preparing the cationic polyester of the present application, comprising adding each monomer and a catalyst into a solvent, sequentially carrying out a first stage polymerization reaction and a second stage polymerization reaction in an inert atmosphere, removing the catalyst after the reaction is completed to obtain a hyperbranched polymer; then, under the action of a coupling agent, end group modification is carried out on the hyperbranched polymer using an end group modification compound E to obtain a cationic polyester with an end group modified with a group E; wherein the conditions for the first stage polymerization reaction are a temperature of 85 to 95° C., a reaction vacuum of 50 to 1000 mbar, and a reaction time of 12 to 24 h; and the conditions for the second stage polymerization reaction are a temperature of 85 to 95° C., a reaction vacuum of 1 to 30 mbar, and a reaction time of 12 to 72 h.

In one implementation of the present application, the molar ratio of monomer P to monomer S is 0.1:10 to 4:1, the molar ratio of monomer M to monomer S is 0:10 to 20:10, and the molar ratio of monomer T to monomer S is 0.1:10 to 20:10.

In one implementation of the present application, the catalyst is immobilized lipase.

In one implementation of the present application, the amount of immobilized lipase used is 3-50 wt % of the total mass of each monomer.

In one implementation of the present application, the solvent is at least one of diphenyl ether, n-dodecane, 1-butyl-3-methylimidazolium hexafluorophosphate, dimethylacetamide and o-phenylenedimethyl ether.

In one implementation of the present application, the amount of the solvent used is 100-500 wt % of the total mass of each monomer.

In one implementation of the present application, removing the catalyst specifically includes filtering the reaction solution with a filtering device after the reaction is completed and collecting the filtrate; obtaining the hyperbranched polymer specifically includes adding n-hexane to the filtrate to precipitate the hyperbranched polymer, centrifuging, removing the supernatant, adding dichloromethane to the precipitate to dissolve it, then adding n-hexane to precipitate the hyperbranched polymer, centrifuging, removing the supernatant, repeating the dissolution with dichloromethane, precipitation with n-hexane, and centrifugation at least twice; finally, drying the precipitate to obtain the hyperbranched polymer.

In one implementation of the present application, the coupling agent is at least one of N,N’-carbonyldiimidazole, carbodiimide, phosphonium cation and urea cation.

In one implementation of the present application, carbodiimides include but are not limited to diisopropylcarbodiimide, dicyclohexylcarbodiimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide.

In one implementation of the present application, the phosphonium cations include but are not limited to benzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate and benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate; the uronium cations include but are not limited to 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate and O-benzotriazole-N,N,N',N'-tetramethyluronium tetrafluoroborate.

In one implementation of the present application, the molar ratio of the coupling agent to the hyperbranched polymer is 2:1 to 50:1; preferably, the molar ratio of the coupling agent to the hyperbranched polymer is 4:1 to 15:1; in addition, the molar ratio of the coupling agent to the end group modified compound E is 5:1 to 1:10.

In one implementation of the present application, the end group of a hyperbranched polymer is modified by using an end group modification compound E, specifically comprising: dissolving the hyperbranched polymer in dichloromethane, adding a coupling agent, stirring at room temperature for at least 10 hours in an inert atmosphere, concentrating the reaction solution, adding at least 3 volumes of ether, centrifuging, removing the precipitate, and obtaining a supernatant; removing the solvent from the supernatant, adding the dried product to dichloromethane to dissolve, adding the end group modification compound E under stirring, and reacting at room temperature for at least 10 hours to obtain a hyperbranched polymer with an end group modified with group E, i.e., the cationic polyester of the present application.

In one implementation of the present application, the preparation method of the present application further includes, after the end group modified compound E is added and the room temperature reaction is completed, adding an equal volume of deionized water to the reaction solution, vortexing, centrifuging and stratifying, and then removing the upper aqueous phase; repeating the operations of adding an equal volume of deionized water, vortexing, centrifuging and stratifying, and removing the upper aqueous phase at least 3 times; finally, adding at least 3 times the volume of n-hexane, vortexing, centrifuging, removing the supernatant, and drying the precipitate, thereby obtaining a hyperbranched polymer with a terminal group modified with group E, that is, the cationic polyester of the present application.

The third aspect of the present application discloses the use of the cationic polyester of the present application in nucleic acid drug delivery.

In one implementation of the present application, nucleic acid drugs include but are not limited to mRNA, circular RNA, siRNA, microRNA, saRNA and DNA.

It should be noted that the key to this application is that a new nucleic acid drug delivery material, namely the cationic polyester of this application, has been discovered; the use of the cationic polyester of this application for nucleic acid delivery not only has high transfection efficiency, but also has low cytotoxicity, and can better meet the needs of clinical use. It can be understood that the cationic polyester of this application can be used not only for nucleic acid delivery, but also for other similar operations that require the delivery of substances into cells due to its low cytotoxicity.

In one implementation of the present application, the nucleic acid drug delivery includes encapsulating the nucleic acid drug with the cationic polyester and delivering it into cells.

In one implementation of the present application, the types of cells include but are not limited to HEK293T, A549, HeLa, U87, HUVEC, Jurkat, RAW264.7, iPSC and MSC.

The fourth aspect of the present application discloses a nucleic acid delivery particle, comprising a wrapping material and a nucleic acid wrapped in the wrapping material; the wrapping material is the cationic polyester of the present application or the wrapping material at least contains the cationic polyester of the present application; the nucleic acid is at least one of mRNA, circular RNA, siRNA, microRNA, saRNA and DNA.

It should be noted that the nucleic acid delivery particles of the present application, due to the use of the cationic polyester of the present application, can not only improve the transfection efficiency, but also have less cytotoxicity and better clinical application prospects. It can be understood that the key to the nucleic acid delivery particles of the present application is the use of the cationic polyester of the present application, and the specific embedding method and nucleic acid delivery method can refer to the prior art.

It should also be noted that the wrapping material of the present application may include other materials besides the cationic polyester of the present application according to different designs or requirements of the wrapping layer, which are not specifically limited here. It can be understood that as long as the cationic polyester of the present application is contained, the transfection efficiency can be improved and the cytotoxicity can be reduced to a certain extent.

In one implementation of the present application, the particle size of the nucleic acid delivery particle is 30-500 nm.

The fifth aspect of the present application discloses a kit for nucleic acid drug delivery, comprising at least one of the following components:

(a) the cationic polyester of the present application;

(b) The nucleic acid delivery particle of the present application.

It should be noted that the kit for nucleic acid drug delivery of the present application can be a kit that has been embedded with a certain specific nucleic acid drug, or it can be the cationic polyester of the present application. The user can design and embed the corresponding nucleic acid drug according to the needs. It can be understood that the key to the kit of the present application is that it contains the cationic polyester or nucleic acid delivery particles of the present application. As for other conventional reagents required for nucleic acid delivery, such as some lipids or polymer materials, etc., they can be obtained by referring to the existing technology or commercially purchasing. Of course, for ease of use, some reagents can also be combined into the kit of the present application, which is not specifically limited here.

The sixth aspect of the present application discloses a method for improving transfection efficiency in nucleic acid drug delivery, comprising using the cationic polyester of the present application or a packaging material containing the cationic polyester of the present application to wrap the nucleic acid drug to form nucleic acid delivery particles, and using the nucleic acid delivery particles to perform cell transfection of the nucleic acid drug.

It should be noted that the method of the present application is key to improving the transfection efficiency by using the cationic polyester of the present application; in one implementation of the present application, the mRNA delivery efficiency of the cationic polyester of the present application is significantly better than the original linear polymer and the best existing commercial mRNA transfection reagent, and the cytotoxicity is lower. In addition, the cationic polyester of the present application is soluble in ethanol and can be better used in clinical practice, with great practical application prospects. It can be understood that the method of improving the transfection efficiency of the present application is key to using the cationic polyester of the present application. As for other operations and reagents for cell transfection, reference can be made to the prior art and no specific limitation is made here.

It should also be noted that the method for improving transfection efficiency of the present application can directly use the cationic polyester of the present application as a wrapping material, or add the cationic polyester of the present application to the existing wrapping material; it can be understood that as long as the cationic polyester of the present application is contained, the transfection efficiency can be improved to varying degrees.

The seventh aspect of the present application discloses the use of the cationic polyester of the present application, or the nucleic acid delivery particle of the present application, or the kit of the present application in disease treatment based on nucleic acid drug delivery.

The eighth aspect of the present application discloses a method for administering a nucleic acid drug in vivo, comprising using the cationic polyester of the present application or a packaging material containing the cationic polyester of the present application to wrap the nucleic acid drug to form nucleic acid delivery particles, and using the nucleic acid delivery particles to deliver the nucleic acid drug; or, directly using the nucleic acid delivery particles of the present application to deliver the nucleic acid drug.

It should be noted that the key to the administration method of the present application is the use of cationic polyesters or nucleic acid delivery particles for nucleic acid drug delivery. As for the specific operation of in vivo delivery and other auxiliary materials required, reference can be made to the prior art and no specific limitation is made here.

Due to the adoption of the above technical solution, the beneficial effects of this application are:

The cationic polyester of the present application adds a monomer T to the three monomers of monomer P, monomer S and monomer M, or two monomers of monomer P and monomer S, and obtains a new polymer with a hyperbranched structure by polymerizing monomer T with other monomers. The cationic polyester of the present application is soluble in ethanol, and when used for nucleic acid delivery, it can not only significantly improve the transfection efficiency, but also has lower cytotoxicity, and can better meet the needs of clinical use.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG1 is a synthetic route diagram of a hyperbranched polymer and terminal group E modification in an embodiment of the present application;

FIG2 is a spectrum of HBPA-E14-3 before and after end group modification in the example of the present application;

FIG3 is a GPC spectrum of HBPA-E14-1 and PACA-E14 in the examples of the present application;

FIG4 is a quantitative graph of the end group modified E14 of HBPA-E14-1 and PACA-E14 in the examples of the present application;

FIG5 is a particle size test result of the complex of HBPA-E14-2 and mRNA in the example of the present application;

FIG6 is a test result of mRNA transfection efficiency of different HBPA-E in A549 and HEK 293T cells in the examples of the present application;

FIG7 is a test result of mRNA transfection efficiency of HBPA-E14-3 modified by two different modification methods, CDI and DIC, and under different mass ratio conditions in the embodiment of the present application;

FIG8 is a test result of DNA transfection efficiency of different HBPA-E in HEK 293T cells in the examples of the present application;

FIG9 is a test result of mRNA transfection efficiency of HBPA-E modified with 26 different end groups E in A549 cells in the embodiment of the present application;

FIG10 is a toxicity test result of the complex of the polymer and mRNA in the example of the present application in the A549 cell model;

FIG. 11 is a test result of the mRNA transfection effect of HBPA-E in vivo in an example of the present application.

Detailed ways

The existing commercial gene delivery materials have low transfection efficiency and high toxicity, which greatly limits the widespread application of gene therapy based on nucleic acid drug delivery. Although studies have reported that biodegradable linear polymers obtained by polymerization of three monomers P, S, and M catalyzed by immobilized lipase based on in vitro synthetic biology technology have higher transfection efficiency and lower cytotoxicity; however, the transfection efficiency of the linear polymer is still not ideal, and the prepared gene delivery materials are difficult to use in actual clinical practice because they need to be dissolved in toxic organic solvents.

The present application creatively adds a new monomer T to the polymerization of the original three monomers, or uses monomer T to replace monomer M, so as to prepare a new cationic polyester with a hyperbranched structure. Specifically, the cationic polyester of the present application is a hyperbranched polymer formed by polymerization of three monomers, monomer P, monomer S and monomer M, or two monomers, monomer P and monomer S, with monomer T, and the end group of the hyperbranched polymer is modified with a group E; monomer P is a cyclic lactone; monomer S is an organic acid with two or more carboxyl groups at the end group; monomer M is a compound with two hydroxyl groups and one secondary or tertiary amine at the end group; monomer T is a compound containing three or more hydroxyl groups; the end group modified compound E provided with the group E modification is a compound containing at least one primary amine, secondary amine or tertiary amine group.

The cationic polyester of the present application has significantly improved gene transfection efficiency, significantly better than the original linear polymer and the best commercial mRNA transfection reagent, and has lower cytotoxicity. More importantly, the cationic polyester of the present application is soluble in ethanol and does not need to be dissolved in toxic organic solvents. It is easy to scale up industrially and can better meet the needs of clinical use, and has great practical application prospects.

The present application is further described in detail below through specific examples. The following examples are only used to further illustrate the present application and should not be construed as limiting the present application.

Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art.

Example

1. Synthesis of hyperbranched polymer (HBPA) and terminal group E modification (HBPA-E)

(1) HBPA synthesis

In this example, a hyperbranched polymer (HBPA) is first formed by polymerizing four monomers P, S, M and T, and then the end group E of HBPA is modified to obtain HBPA-E. Specifically, the monomer P in this example is cyclopentadecanolide (PDL), the monomer S is sebacic acid (SA), the monomer M is methyldiethanolamine (MDEA), and the monomer T is triethanolamine (TEA). The HBPA synthesis route is shown in Figure 1 (a). This example designs seven experiments and two controls. According to the molar ratio of the four monomers P (PDL) / S (SA) / M (MDEA) / T (TEA) in Table 1, the raw materials are added to a round-bottom flask, and immobilized lipase (CALB) is added at 10 wt% of the total raw material mass, and finally 200 wt% of diphenyl ether is added as raw material. After the reaction system is replaced with argon three times, it is stirred and heated to 90°C at 60 mbar for 24 hours, and then the reaction is continued at 2.1 mbar for 48 hours. After the reaction is completed, the reaction solution is filtered to remove the lipase. The filtrate was added with n-hexane, vortexed and centrifuged, and the supernatant was discarded. The precipitate was dissolved with dichloromethane, and then n-hexane was added to precipitate it, centrifuged, and the supernatant was discarded. The above operation was repeated 3 times. The precipitate was vacuum dried for one day to obtain HBPA. Among them, the two control synthesized polymers were linear polymers PACA.

Table 1 Characterization of polymers synthesized with different feed ratios and their modified E14 products

In Table 1, "a" is calculated based on 1H-NMR results, and "b" is calculated based on GPC results.

(2) Modification of terminal group E

In this example, as shown in Table 1, the terminal group-modified compound E14 was used, and two different terminal group-modification methods were used, namely, N,N'-carbonyldiimidazole (CDI) and diisopropylcarbodiimide (DIC) were used as coupling agents to modify the terminal groups of the polymer.

Modification method 1: Use CDI as a coupling agent. The synthetic route is shown in Figure 1 (b). Take 250 mg HBPA and dissolve it in 5 mL of ultra-dry dichloromethane (DCM). Add 40 eq of CDI under stirring. After replacing the reaction system with argon three times, stir at room temperature overnight. Concentrate the reaction solution to 3 mL, add 3 times the volume of ether, vortex, centrifuge, and remove the precipitate. The supernatant is decompressed and dried, ultra-dry DCM is added to dissolve, and then 40 eq of E14 is added under stirring and reacted at room temperature for 24 hours. After the reaction is completed, add an equal volume of deionized water to the reaction solution, vortex, centrifuge and separate the layers, and remove the upper aqueous phase. Add an equal volume of deionized water and repeat the above operation 5 times. Add 3 times the volume of n-hexane to the lower layer of dichloromethane solution, vortex and centrifuge. The precipitate is vacuum dried for one day to obtain HBPA-E14. For control 1, PACA-E14 is obtained.

Modification method 2: Use DIC as a coupling agent. The synthesis route is shown in Figure 1 (c). Take 250 mg HBPA and dissolve it in 5 mL ultra-dry DCM. Add 40 eq of DIC and 40 eq of E14 while stirring. After the reaction system is replaced with argon three times, stir at room temperature overnight. After the reaction is completed, add an equal volume of deionized water to the reaction solution, vortex and centrifuge, and remove the upper aqueous phase. Add another equal volume of deionized water and repeat the above operation three times. Then add 3 times the volume of n-hexane to the lower layer of dichloromethane, vortex and centrifuge. Wash the precipitate with a small amount of ethanol and then vacuum dry it for one day to obtain HBPA-E14, such as HBPA-E14-3-DIC prepared in Experiment 4. For control 2, PACA-E14-DIC is obtained.

Using a method similar to that of Experiment 3, this example further used 25 terminal group modification compounds from E1 to E26, except E14, to modify the terminal groups of HBPA to synthesize other HBPA-E.

The 26 terminal group modified compounds from E1 to E26 are as follows:

2. Structural Characterization of HBPA and HBPA-E

The molecular structures of HBPA and HBPA-E can be characterized by 1H-NMR. The results are shown in Table 1, and some of the results are shown in Figure 2. Figure 2 is the spectrum of HBPA in Experiment 2 before and after end group modification, that is, the 1H-NMR spectrum of HBPA-3 and HBPA-E14-3, and the solvent is CDCl _3. In the spectrum, the characteristic peak of the PDL unit is f (δ≈4.08ppm), the characteristic peak of the SA unit is b (δ≈1.60ppm), the characteristic peak of the MDEA unit is g (δ≈4.20ppm), and the characteristic peak of the TEA unit is h (δ≈2.85ppm). The presence of the TEA unit indicates that the polymer structure is in line with the expected assumptions and may have a hyperbranched structure, while the characteristic peak k (δ≈2.22ppm) can prove that the end group has been successfully modified with group E. The ratio of each repeating unit of the polymer can be converted by calculating the integrated area of each characteristic peak. The results show that the molar ratio of P/S/M/T repeating units of HBPA-E14-3 obtained by CDI modification is 1:9:4.5:1.9, which is very close to the feed ratio of 1:9:6:2. This shows that the reaction of each monomer raw material in the reaction is basically complete and the reaction efficiency is very high. Other 1H-NMR results are shown in Table 1. The results show that even with the continuous increase of T feed, the enzyme-catalyzed reaction is close to complete, and the reaction can be very controllable to obtain hyperbranched polymers without cross-linking.

The experiment showed that the reduction of monomer M had little effect on the hyperbranched polymer; therefore, this example further studied the case where the proportion of monomer M was reduced to 0, that is, the polymerization reaction was directly carried out using monomers P, S and T. Specifically, the molar ratio of the feed was P:S:T = 1:9:6, and the remaining steps and parameters were the same as "(1) HBPA synthesis". The results showed that the three monomers P, S and T can also obtain a polymer with a hyperbranched structure without cross-linking. The hyperbranched polymer obtained from the three monomers P, S and T has similar physical and chemical properties to the hyperbranched polymer obtained from the four monomers P, S, M and T.

At the same time, GPC was used to characterize the molecular weight and distribution of HBPA-E. GPC measurements were performed at 35°C using a Waters 1515 column and a 2414 refractive index (RI) detector. The mobile phase was DMF containing 0.1% LiBr, with a flow rate of 1 mL/min, and a linear polymethyl methacrylate standard was used as a standard. The results are shown in Table 1, and some of the results are shown in Figure 3. Figure 3 shows the GPC spectra of HBPA-E14-1 of Experiment 3 and PACA-E14 of Control 1, respectively. The results in Figure 3 show that the GPC spectra of hyperbranched HBPA-E14-1 and linear PACA-E14 are both single peaks, with number average molecular weights _Mn of 8296 and 8769 Da, respectively, and PDIs of 4.12 and 1.97, respectively. The molecular weights of the two are close, but the PDIs are significantly different. This indicates that there are significant structural differences between HBPA-E14-1 and PACA-E14, which should be derived from the branched structure of HBPA. The PDIs of other HBPA-E14 are shown in Table 1, which are similar to those of HBPA-E14-1.

In addition, in this example, HBPA-E14 and PACA-E14 prepared in Table 1 were dissolved in ethanol to test their solubility. The results are shown in Table 1. The results in Table 1 show that the branched HBPA-E in this example is easily soluble in ethanol, with a solubility greater than 25 mg/mL, while the linear PACA-E is poorly soluble in ethanol, with a solubility less than 10 mg/mL.

Finally, in this example, the content of E14 modified by the end group of HBPA-E14 (Mn=8769) and PACA-E14 (Mn=8296) with similar number average molecular weight was quantified by the ninhydrin method. The specific method is as follows: 1. Preparation of ninhydrin colorimetric agent: weigh 200 mg of ninhydrin, dissolve in 9 mL of ethanol, add ethanol to make up to 10 mL, and prepare a 20 mg/mL ninhydrin ethanol solution. 2. Sample determination: take about 10 mg of HBPA-E14 and PACA-E14 samples, prepare a 5 mg/mL ethanol solution, take 100 μL of each and mix them evenly with 200 μL of ninhydrin ethanol solution, place on a shaker and heat and shake at 60°C for 10 minutes, cool to room temperature, take 200 μL of each to measure its absorbance at 570 nm, and convert its absorbance value into the relative concentration of the end group E14. The results are shown in Figure 4. The relative concentration of end group E14 in PACA-E14 is 1, while at the same material concentration, the relative concentration of end group E14 in HBPA-E14-1 is 3.01, indicating that its end group E14 content is nearly 3 times that of PACA-E14. Considering that the molecular weights of the two are very close, the significantly higher end group content of HBPA-E14-1 than that of PACA-E14 should be due to its branched structure, which once again verifies the branched structure of HBPA.

3. Characterization of Materials and mRNA Complexes

After the polymer material HBPA-E14-2 was prepared, it was mixed with Firefly Luciferase mRNA (N1-Me-Pseudo UTP, Nanjing Novozyme Biotechnology Co., Ltd.) at different mass ratios and the diameter of the complex was measured. The complex of PACA-E14 and mRNA was also used as a control. Specifically, the polymer materials HBPA-E14-2 and PACA-E14 were dissolved in ethanol and DMSO respectively to prepare a 25 mg/mL polymer solution, and then different amounts of the polymer solution were added to a certain volume of pH 4.9 NaAc buffer and vortexed. At the same time, a pH 4.9 NaAc solution containing 20 μg/mL mRNA was prepared, and an equal volume of the polymer solution was added to the mRNA solution and vortexed to finally obtain a complex solution with a polymer and mRNA mass ratio of 25:1, 50:1, 75:1, 100:1, 150:1, and 200:1. HBPA-E14-2 and PACA-E14 were subjected to the above mass ratio control test. The dosage of HBPA-E14-2 and PACA-E14 was calculated based on the mass ratio. For example, if the mass ratio of polymer to mRNA was 25:1, 20 μL of 25 mg/mL polymer solution, i.e. 500 μg of polymer, was added to 980 μL of pH 4.9 NaAc buffer and vortexed. At the same time, a pH 4.9 NaAc solution containing 20 μg/mL mRNA was prepared, and an equal volume of polymer solution was added to the mRNA solution and vortexed to obtain a mass ratio of polymer to mRNA of 25:1.

The particle size of the complex was tested by a particle size analyzer (Malvern Panalytical Zetasizer Pro) at 25°C. The results are shown in Figure 5, where the horizontal axis of Figure 5 is the mass ratio of polymer to mRNA, and the vertical axis is the particle size of the complex, in nm.

The results in Figure 5 show that the diameter of the HBPA-E14-2/mRNA complex exceeds 300 nm when the mass ratio is 25:1, while the diameter decreases significantly and gradually stabilizes at 100-200 nm when the mass ratio is 50:1-200:1. This indicates that when the mass ratio is greater than 50:1, the complex of HBPA-E14-2 and mRNA tends to be stable. PACA-E14 and mRNA also show similar results.

IV. Test of mRNA and DNA transfection efficiency of HBPA-E in vitro cells

After obtaining the complex of HBPA-E and mRNA or DNA expressing luciferase (Firefly Luciferase DNA, Guangzhou Aiji Biotechnology Co., Ltd.), this example tests its transfection efficiency in different cell models. The specific method of cell transfection is as follows: A549 and HEK293T cells (ATCC) are inoculated in a 48-well cell culture plate (2.5×10 ⁴ /well), and the cells are cultured in DMEM at 37°C and 5% CO ₂ for 12 hours to adhere to the wall. Then, the DMEM medium (250 μL/well) is replaced, and the sample to be tested and the positive control are added and cultured for 24 hours (mRNA) or 48 hours (DNA), and the sample is added according to the final 2 μg/mL mRNA per well. The preparation method of the complex of the test material and mRNA or DNA is the same as "III. Characterization of the material and mRNA complex". The positive control for mRNA transfection used Thermo Fisher's Lipofectamine MessengerMAX (LipoMM), and the positive control for DNA transfection used Lipofectamine 2000 (Lipo2k).

The specific method of testing the lysis and luminescence effect after cell transfection is as follows: 24 hours (mRNA) or 48 hours (DNA) after cell transfection, remove the culture medium in the well plate and place it in a -80℃ refrigerator for 10 minutes. Take out the well plate and place it at 4℃, take 40μL of lysis solution to the 48-well plate, cover the bottom surface, let it stand for 5 minutes, then take 280μL of test solution to the 48-well plate, then take 160μL of the mixture to the black microplate reader plate, and finally use a spray gun to take 40μL D-Luciferin and add it to each well. After reacting at room temperature for 2 minutes, put it into the microplate reader to test the luminescence at 560nm.

The test results of mRNA and DNA transfection efficiency of different HBPA-E in A549 and HEK 293T cells are shown in Figures 6 to 9. The specific experimental designs are as follows:

In this example, the mRNA transfection efficiency in A549 and HEK293T cells was tested after the polymers of Control 1, Test 1, Test 2, Test 3, Test 5, Test 6 and Test 7 were respectively complexed with mRNA at a polymer:mRNA mass ratio of 75:1, and the results are shown in Figure 6.

The results in Figure 6 show that in A549 cells, all hyperbranched materials showed significantly higher mRNA transfection effects than the commercial control LipoMM, and much higher than the transfection effect of linear PACA-E14. Among them, HBPA-E14-3 had the highest transfection efficiency, nearly 4 times that of LipoMM. The linear PACA-E14 was weaker than LipoMM. The transfection results of HEK293T cells were consistent with those of A549. This shows that the cell transfection effect of hyperbranched polymers on mRNA is significantly improved compared with linear polymers, and is also better than the most efficient commercial transfection reagents currently available. This effect has also been verified on different cells. Considering that this hyperbranched polymer can also be dissolved in ethanol, it has great application potential in both in vitro and in vivo mRNA transfection.

At the same time, this example also tested the mRNA transfection efficiency of HBPA-E14-3 modified by two different modification methods, CDI and DIC, and under different mass ratio conditions. Specifically, this example tested the transfection efficiency of HBPA-E14-3 modified with CDI, i.e., the polymer in Experiment 3 and mRNA complexed at a polymer:mRNA mass ratio of 25:1, 50:1, 75:1, and 100:1 in A549, and the transfection efficiency of HBPA-E14-3-DIC modified with DIC, i.e., the polymer in Experiment 4 and mRNA complexed at a polymer:mRNA mass ratio of 25:1, 50:1, 75:1, and 100:1 in A549; the results are shown in Figure 7.

The results in Figure 7 show that on A549 cells, HBPA-E14-3 modified by two methods has a significantly higher transfection efficiency than LipoMM at a mass ratio of 50:1 to 100:1, but the material modified by CDI is better than the material modified by DIC. However, no transfection efficiency was shown at a mass ratio of 25:1. These results show that both modification methods are feasible. At the same time, mRNA is not fully complexed at 25:1, but it is already complexed when it is higher than 50:1, which is consistent with the test results of the complex particle size tested by the particle size analyzer.

In addition, this example also tested the DNA transfection efficiency of different HBPA-E in HEK 293T cells. Specifically, the DNA transfection efficiency in HEK 293T cells was tested after the polymers of Control 1, Test 1, Test 2, Test 3, Test 5, Test 6 and Test 7 were complexed with DNA at a polymer:DNA mass ratio of 75:1. The results are shown in Figure 8.

The results in Figure 8 show that all hyperbranched materials exhibit significantly higher DNA transfection effects than the commercial control Lipo2k, and much higher than the transfection effect of linear PACA-E14, among which HBPA-E14-2 has the highest transfection efficiency. This shows that hyperbranched polymer materials can also be used for DNA transfection.

Finally, this example also tested the mRNA transfection efficiency of HBPA-E modified with 26 different terminal modification compounds with group E in A549 cells. Specifically, the mRNA transfection efficiency of 26 HBPA-E modified with 26 terminal modification compounds E1 to E26 and complexed with mRNA at a polymer:mRNA mass ratio of 75:1 in A549 cells was tested. The results are shown in Figure 9.

The results in Figure 9 show that different end group modifications have an effect on the mRNA transfection efficiency, among which HBPA-E with various end group modifications showed significantly higher transfection efficiency than LipoMM, such as E2, E4, E9, E10, E12, E14, E15, E25 and E26. Except that the mRNA transfection efficiency of polymers modified with E11, E13, E22, E23 and E24 was significantly lower than that of LipoMM, the mRNA transfection efficiency of polymers modified with E8, E17, E18 and E21 was comparable to that of LipoMM, and the mRNA transfection efficiency of polymers modified with other end groups was higher than that of LipoMM.

5. Cytotoxicity Test

In this example, the cytotoxicity of the HBPA-E/mRNA complex was tested in the A549 cell model. The specific method of the cytotoxicity test is as follows: A549 cells were inoculated in a 96-well cell culture plate (1×10 ⁴ /well). After the cells were cultured for 12 hours to adhere to the wall, the DMEM culture medium was replaced, and the mRNA complexes of the test samples PACA-E14, HBPA-E14-3 and the positive control LipoMM were added and cultured for 24 hours, respectively, wherein the mass ratio of PACA-E14 or HBPA-E14-3 to mRNA was 50:1. Samples were added according to the final mRNA concentration in each well of the culture medium of 0.125, 0.25, 0.5, 1, 2, 4 and 8 μg/mL in sequence. After 24 hours, 10 μL of CCK8 detection reagent was added to each well, and the absorption at 450nm was detected by an enzyme marker. The results are shown in Figure 10.

The results of Figure 10 show that the control LipoMM complex exhibits significant cytotoxicity, with a cell survival rate of only 50% at an mRNA concentration of 2 μg/mL, and the toxicity increases significantly with increasing mRNA concentration. In comparison, the toxicity of the linear PACA-E14 complex is significantly reduced, but it also exhibits certain cytotoxicity, with a cell survival rate of 70% at an mRNA concentration of 2 μg/mL. Compared with these two, the toxicity of the hyperbranched HBPA-E14-3 complex is further reduced, with a cell survival rate of more than 80% at an mRNA concentration of 2 μg/mL. This result shows that hyperbranched HBPA-E14-3 has higher biocompatibility than commercial transfection reagents and linear PACA-E14, and also has greater prospects for in vivo applications.

6. Test of mRNA transfection effect of HBPA-E in vivo

This example tests the mRNA transfection effect of HBPA-E14-3 in vivo. The specific method is as follows: 4-6 week old C57 mice (weighing about 20g) were purchased from Guangdong Medical Experimental Animal Center and monitored and raised in an SPF (specific pathogen-free) environment. The HBPA-E14-3/mRNA complex was injected into the mouse body through pulmonary administration (administered by inserting a pulmonary spray needle into the trachea, 5μg dose) and intratracheal instillation (5μg dose), and its luciferase expression effect in vivo was observed, with normal saline as a blank control. The results are shown in Figure 10. Among them, in the HBPA-E14-3/mRNA complex, the mass ratio of HBPA-E14-3 to mRNA is 50:1.

In Figure 11, the horizontal axis "Control" represents the blank group, "Pulmonary administration of mRNA (5 μg)" represents direct pulmonary administration of mRNA, "Pulmonary administration (5 μg)" represents pulmonary administration using HBPA-E14-3/mRNA complex, and "Intratracheal instillation (5 μg)" represents intratracheal instillation administration using HBPA-E14-3/mRNA complex.

The results in Figure 11 show that no fluorescence was detected in the direct mRNA pulmonary administration and the blank group, while the HBPA-E14-3/mRNA complex showed significant luciferase expression effects in different administration routes, among which the pulmonary administration was twice as efficient as the intratracheal instillation. These results indicate that this hyperbranched polymer/mRNA complex can achieve mRNA delivery in vivo and has great prospects for in vivo applications.

The above contents are further detailed descriptions of the present application in combination with specific implementation methods, and it cannot be determined that the specific implementation of the present application is limited to these descriptions. For ordinary technicians in the technical field to which the present application belongs, several simple deductions or substitutions can be made without departing from the concept of the present application.

Claims (21)

1. A cationic polyester, characterized in that: the cationic polyester is a hyperbranched polymer formed by polymerization of three monomers, monomer P, monomer S and monomer M, or two monomers, monomer P and monomer S, and monomer T, and the end group of the hyperbranched polymer is modified with a group E;

   Wherein, monomer P is a cyclic lactone;

   Monomer S is an organic acid having two or more carboxyl groups at the terminal end;

   The monomer M is a compound containing two hydroxyl groups and a secondary or tertiary amine;

   Monomer T is a compound containing three or more hydroxyl groups;

   The terminal group modification compound E providing the modification of the group E is a compound containing at least one primary amine, secondary amine or tertiary amine group.
2. The cationic polyester according to claim 1, characterized in that: the monomer P is a cyclic lactone with a fatty chain length of 6 to 35;

   Preferably, the monomer P is at least one of cyclohexyl lactone, cyclododecanolide, cyclopentadecanolide and cyclohexadecanolide.
3. The cationic polyester according to claim 1, characterized in that: the monomer S is an organic acid with a carbon chain length of 3 to 18;

   Preferably, monomer S is at least one of adipic acid, sebacic acid and 1,2,3-propanetricarboxylic acid.
4. The cationic polyester according to claim 1, characterized in that: the monomer M is a compound with a carbon chain length of 4 to 36;

   Preferably, the monomer M is at least one of diethanolamine, methyldiethanolamine and ethyldiethanolamine.
5. The cationic polyester according to claim 1, characterized in that: monomer T is a compound with a carbon chain length of 4 to 54;

   Preferably, monomer T is at least one of trimethylolpropane, 3-(hydroxymethyl)-1,5-pentanediol, triethanolamine, and N,N,N',N'-tetrahydroxyethylethylenediamine.
6. The cationic polyester according to claim 1, characterized in that: the terminal group modified compound E providing the group E modification is at least one of E1 to E26;

   

   Preferably, the terminal modified compound E providing group modification is at least one of E1 to E10, E12, E14 to E21, E25, and E26;

   More preferably, the terminal modified compound E providing group modification is at least one of E2, E4, E9, E10, E12, E14, E15, E25, and E26.
7. The cationic polyester according to any one of claims 1 to 6, characterized in that: the cationic polyester is a hyperbranched polymer with a structure shown in Formula 1;

   Formula 1

   

   Wherein, x, y, and z are independent integers ranging from 1 to 200.

   n is an integer from 0 to 200,

   j and k are integers from 0 to 30,

   l, m, o, p, q are independent integers from 1 to 20,

   _Rx is hydrogen, or a substituted or unsubstituted alkyl group containing 1 to 18 carbon atoms, or a substituted or unsubstituted aromatic group containing at least one benzene ring, or a substituted or unsubstituted heterocyclic group containing at least one heterocyclic ring, or a substituted or unsubstituted alkoxy group containing 1 to 18 carbon atoms and at least one oxygen atom;

   J is hydrogen, R ₁ is modified by the group E provided by the terminal group modification compound E, and there is no R ₂ ; or, J is a carbonyl group, and both R ₁ and R ₂ are modified by the group E provided by the terminal group modification compound E;

   In Formula 1, the truncation line represents a branch structure, the first monomer connected to the branch structure is P or S, and other monomers are subsequently connected to form a hyperbranched structure.
8. The cationic polyester according to any one of claims 1 to 6, characterized in that the number average molecular weight of the cationic polyester is 1-30k, preferably 2-20k.
9. The method for preparing a cationic polyester according to any one of claims 1 to 8, characterized in that: the method comprises adding each monomer and a catalyst into a solvent, sequentially carrying out a first stage polymerization reaction and a second stage polymerization reaction in an inert atmosphere, and after the reaction is completed, removing the catalyst to obtain a hyperbranched polymer; then, under the action of a coupling agent, using an end group modification compound E to modify the end group of the hyperbranched polymer to obtain a cationic polyester with an end group modified with a group E;

   The conditions of the first stage polymerization reaction are: temperature 85-95°C, reaction vacuum degree 50-1000 mbar, reaction time 12-24 hours;

   The conditions of the second stage polymerization reaction are: temperature 85-95° C., reaction vacuum degree 1-30 mbar, and reaction time 12-72 h.
10. The preparation method according to claim 9, characterized in that: the molar ratio of monomer P to monomer S is 0.1:10 to 4:1, the molar ratio of monomer M to monomer S is 0:10 to 20:10, and the molar ratio of monomer T to monomer S is 0.1:10 to 20:10;

    Preferably, the catalyst is immobilized lipase;

    Preferably, the amount of the immobilized lipase is 3-50wt% of the total mass of each monomer;

    Preferably, the solvent is at least one of diphenyl ether, n-dodecane, 1-butyl-3-methylimidazolium hexafluorophosphate, dimethylacetamide and o-phenylenedimethyl ether;

    Preferably, the amount of the solvent is 100-500wt% of the total mass of each monomer;

    Preferably, the catalyst removal specifically comprises: filtering the reaction solution with a filtering device after the reaction is completed, and collecting the filtrate; the hyperbranched polymer is obtained specifically comprising: adding n-hexane to the filtrate to precipitate the hyperbranched polymer, centrifuging, removing the supernatant, adding dichloromethane to dissolve the precipitate, then adding n-hexane to precipitate the hyperbranched polymer, centrifuging, removing the supernatant, repeating the dichloromethane dissolution, n-hexane precipitation, and centrifugation at least twice; finally, drying the precipitate to obtain the hyperbranched polymer.
11. The preparation method according to claim 9, characterized in that: the coupling agent is at least one of N,N'-carbonyldiimidazole, carbodiimide, phosphorus cation and urea cation;

    Preferably, the carbodiimides include but are not limited to diisopropylcarbodiimide, dicyclohexylcarbodiimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide;

    The phosphonium cations include, but are not limited to, benzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate and benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate;

    The uronium cations include, but are not limited to, 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate and O-benzotriazole-N,N,N',N'-tetramethyluronium tetrafluoroborate;

    Preferably, the molar ratio of the coupling agent to the hyperbranched polymer is 2:1 to 50:1;

    Preferably, the molar ratio of the coupling agent to the hyperbranched polymer is 4:1 to 15:1;

    The molar ratio of the coupling agent to the terminal group modified compound E is 5:1 to 1:10.
12. The preparation method according to any one of claims 9 to 11, characterized in that: the end group modification of the hyperbranched polymer using the end group modification compound E specifically comprises: dissolving the hyperbranched polymer in dichloromethane, adding a coupling agent, stirring at room temperature for at least 10 hours in an inert atmosphere, concentrating the reaction solution, adding at least 3 volumes of ether, centrifuging, removing the precipitate, and obtaining a supernatant; removing the solvent in the supernatant, adding the dried product to dichloromethane to dissolve, adding the end group modification compound E under stirring, reacting at room temperature for at least 10 hours, and obtaining a hyperbranched polymer with a modified end group having a group E, i.e., the cationic polyester;

    Preferably, the method further comprises the following steps: after the end group modified compound E is added to react at room temperature, an equal volume of deionized water is added to the reaction solution, vortexing and centrifuging to separate the layers, and then removing the upper aqueous phase; repeating the steps of adding an equal volume of deionized water, vortexing, centrifuging to separate the layers, and removing the upper aqueous phase at least 3 times; and finally, adding at least 3 times the volume of n-hexane, vortexing, centrifuging, removing the supernatant, and drying the precipitate to obtain a hyperbranched polymer having an end group modified with group E.
13. Use of the cationic polyester according to any one of claims 1 to 8 in nucleic acid drug delivery.
14. The use according to claim 13 is characterized in that: the nucleic acid drug includes but is not limited to mRNA, circular RNA, siRNA, microRNA, saRNA and DNA.
15. The use according to claim 13 or 14, characterized in that: the nucleic acid drug delivery comprises encapsulating the nucleic acid drug with the cationic polyester and delivering it into cells;

    Preferably, the types of cells include but are not limited to HEK293T, A549, HeLa, U87, HUVEC, Jurkat, RAW264.7, iPSC and MSC.
16. A nucleic acid delivery particle, characterized in that it comprises a wrapping material and a nucleic acid wrapped in the wrapping material;

    The wrapping material comprises the cationic polyester according to any one of claims 1 to 8;

    The nucleic acid is at least one of mRNA, circular RNA, siRNA, microRNA, saRNA and DNA.
17. The nucleic acid delivery particle according to claim 16, characterized in that the particle size of the nucleic acid delivery particle is 30-500 nm.
18. A kit for delivering a nucleic acid drug, characterized in that it comprises at least one of the following components:

    (a) the cationic polyester according to any one of claims 1 to 8;

    (b) The nucleic acid delivery particle according to claim 16 or 17.
19. A method for improving transfection efficiency in nucleic acid drug delivery, characterized in that: the method comprises using the cationic polyester described in any one of claims 1 to 8 or a wrapping material containing the cationic polyester described in any one of claims 1 to 8 to wrap the nucleic acid drug to form nucleic acid delivery particles, and using the nucleic acid delivery particles to perform cell transfection on the nucleic acid drug.
20. Use of the cationic polyester according to any one of claims 1 to 8, or the nucleic acid delivery particle according to claim 16 or 17, or the kit according to claim 18 in disease treatment based on nucleic acid drug delivery.
21. A method for administering a nucleic acid drug in vivo, characterized in that: the method comprises: using the cationic polyester described in any one of claims 1 to 8 or a packaging material containing the cationic polyester described in any one of claims 1 to 8 to wrap the nucleic acid drug to form nucleic acid delivery particles, and using the nucleic acid delivery particles to deliver the nucleic acid drug; or, using the nucleic acid delivery particles described in claim 16 or 17 to deliver the nucleic acid drug.

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