CN116640301A - A class of functional highly flexible branched polymers, preparation methods and mRNA delivery applications - Google Patents

Abstract

The invention relates to the technical field of biological high polymer materials, and discloses a functional hyperbranched poly (beta-amino ester), a preparation method and mRNA or DNA delivery application thereof.

Description

A class of functional highly flexible branched polymers, preparation methods and mRNA delivery applications

technical field

The invention belongs to the field of biopolymer materials, and relates to a class of functionally highly flexible branched poly(β-amino esters), a preparation method thereof and mRNA delivery applications.

Background technique

mRNA delivery has shown broad application prospects in the fields of protein replacement therapy and vaccine development. Compared with DNA, mRNA delivery has unique advantages, such as no transcription steps, no need to enter the nucleus, no transgene insertion mutation, and higher protein expression efficiency and kinetics are faster. In addition, the amount of mRNA is low, generally 1/5-1/10 of DNA; currently, liposome nanoparticles are one of the most potential carriers for mRNA delivery, but due to high price, poor serum tolerance, poor stability, The preparation process is complicated and the composition is complicated, which seriously limits its clinical application.

Cationic polymer carriers have many advantages such as wide source of raw materials, flexible and diverse chemical composition, easy to adjust topology, high gene loading efficiency, and good serum tolerance and stability; commonly used cationic polymers such as polyethyleneimine and polydimethylmethacrylate Aminoethyl ester has shown good DNA transfection efficiency, but its poor degradation performance leads to high cytotoxicity, and its clinical safety has been questioned; poly(β-amino ester) as a highly efficient and degradable cationic polymer Carriers have become one of the most promising new polymer carriers at present.

At present, the research on poly(β-amino ester) in gene delivery mainly focuses on DNA delivery, especially highly branched poly(β-amino ester) has shown broad application prospects in DNA delivery in vivo and in vitro, but due to the Due to the differences in DNA structure, composition and transfection barrier, highly branched poly(β-amino ester) does not exhibit excellent mRNA delivery performance, which will greatly limit its application research in gene delivery; therefore, a class of Functional highly flexible branched polymers, preparation methods and mRNA delivery applications.

Contents of the invention

In order to solve the above-mentioned technical problems, the present invention provides a highly flexible branched polymer, a preparation method and mRNA delivery application. By regulating the type of branched monomer and terminal functional modification, a brand-new polymer with multiple terminal groups has been prepared. Highly flexible branched poly(β-amino ester) groups to solve the current problems of low stability of gene delivery.

Concrete technical scheme of the present invention is as follows:

The invention discloses a class of functional highly flexible branched poly(β-amino esters) (HBPAEs). The structural formula of the functional highly branched poly(β-amino esters) (HBPAEs) is as follows:

In the formula, n=10-60, m=10-60;

In the formula, n=10-60, m=10-60.

Preferably, the functional highly flexible branched poly(β-amino ester) has a molecular weight in the range of 4000-40000 Da.

Preferably, the acrylate monomer is 1,4-butanediol diacrylate, trimethylolpropane ethoxylate triacrylate, and pentaerythritol tetraacrylate.

Preferably, the small molecule organic amine is 4-amino-1-butanol, 5-amino-1-pentanol 1,2-ethylenediamine, tris(2-aminoethyl)amine, tris[2- (methylamino)ethyl]amine.

Preferably, the functional end-capping agent monomer is 3,6,9-trioxaundecane-1,11-diamine, 3-methylaminopropylamine, triethylenetetramine, 1-(3- aminopropyl)-4-methylpiperazine.

The present invention also discloses a preparation method of the above-mentioned functional highly branched poly(β-amino ester), comprising the following steps:

S1: A highly branched poly(β-amino ester) with a double bond is prepared by Michael addition reaction of a diacrylate monomer and a small molecule organic amine;

S2: Prepare a functional highly branched poly(β-amino ester) by reacting the highly branched poly(β-amino ester) with a double bond prepared in step S1 with a functional end-capping agent.

Preferably, in step S1, the reaction molar ratio of the diacrylate monomer and the small molecule organic amine is 1:0.5˜1:3.

Preferably, in step S2, the reaction of the functionalized end-capping agent is to combine the highly branched poly(β-amino ester) with double bonds prepared in step 1) and the end-capping agent monomer at a total monomer concentration of 100 -500mg/mL reaction at room temperature for 12h-72h, the reaction molar ratio of capping agent and small molecule organic amine is 1:0.5~1:4.

The present invention also discloses the application of the above-mentioned functional highly branched poly(β-amino ester) as a cationic polymer carrier, and the functional highly flexible branched poly(β-amino ester) is used for mRNA delivery.

Further, the functional highly flexible branched poly(β-amino ester) is used for DNA delivery.

Compared with the prior art, the present invention has the following beneficial effects:

1. The present invention discloses a synthetic method for highly flexible branched poly(β-amino ester), and prepares a brand-new poly(β-amino ester) with multiple terminal groups by regulating the type of branched monomer and the terminal functional modification. Highly flexible branched poly(β-amino ester); this functional highly branched poly(β-amino ester) has a brand-new chemical composition, charge density, biodegradability and good biocompatibility, and the current technology in the art Compared with commercialized mRNA transfection reagent LipofectamineMessengerMAX, the functional highly flexible branched poly(β-amino ester) disclosed by the present invention has more clinical potential; the synthesis method has low cost, simple synthesis route and low equipment requirements , suitable for industrial scale-up production.

2. The present invention discloses the composite process of the above-mentioned functional highly flexible branched poly(β-amino ester) and mRNA, and prepares a A series of composite nanoparticles assembled with highly branched poly(β-amino ester) and mRNA, the composite nanoparticles have smaller size and higher surface positive potential, indicating that the formed composite nanoparticles have excellent stability.

3. The present invention discloses the application of the above-mentioned highly flexible branched poly(β-amino ester), and it is confirmed by experiments that it is a gene delivery carrier with a new structure with high mRNA transfection efficiency and low toxicity. The accuracy and wide applicability of the invention were verified in various tissue cells (in HeLa cells, UC-3 cells, HT-29 cells and HCV-29 cells).

4. The present invention discloses that highly flexible branched poly(β-amino ester) can efficiently mediate DNA encoding green fluorescent protein in various tissue cells, including HeLa cells, IEC-6 cells, HepG2 cells and B16-F10 cells, demonstrating that highly flexible branched poly(β-amino esters) can deliver both DNA and mRNA, thereby demonstrating the broad applicability of the present invention.

Description of drawings

Fig. 1 is the synthesizing schematic diagram of highly flexible branched poly(β-amino ester) of the present invention;

Fig. 2 is the HNMR spectrogram of representative highly flexible branched poly(β-amino ester) of the present invention;

Fig. 3 is the gel permeation chromatogram (GPC) graph after purification of highly flexible branched poly(β-amino ester) of the present invention; Wherein (a) is highly flexible branched poly(β-amino ester) with molecular weight about 20,000Da (b) is a highly flexible branched poly(β-amino ester) with a molecular weight of about 15,000 Da;

Fig. 4 is highly flexible branched poly (beta-amino ester) of the present invention affinity performance test to mRNA, confirms its excellent mRNA affinity performance;

Fig. 5 is the particle diameter diagram of the complex nanoparticle that the highly flexible branched poly(beta-amino ester) of the present invention compresses mRNA;

Fig. 6 is the potential diagram of the composite nanoparticle formed by the highly flexible branched poly(β-amino ester) compressed mRNA of the present invention;

Fig. 7 is the microscopic topography diagram of the composite nanoparticle formed by highly flexible branched poly(β-amino ester) compressed mRNA of the present invention;

Fig. 8 is the fluorescence photo of cells after highly flexible branched poly(β-amino ester) transfection encoding green fluorescent protein (GFP) mRNA in different cells of the present invention, including HeLa cells, UC-3 cells, HT- 29 cell and HCV-29 cell map;

Fig. 9 is the efficiency diagram of highly flexible branched poly(β-amino ester) transfection encoding luciferase (Fluc) mRNA under different mass ratios in UC-3 cells of the present invention;

Fig. 10 is a diagram of the cell survival rate of highly flexible branched poly(β-amino ester) transfected mRNA in different mass ratios in UC-3 cells according to the present invention;

Fig. 11 is a fluorescent photograph of cells transfected with highly flexible branched poly(β-amino ester) DNA encoding green fluorescent protein (GFP) in different cells, including HeLa cells, IEC-6 cells, HepG2 cells and In B16-F10 cells, this demonstrates that highly flexible branched poly(β-amino esters) can deliver both DNA and mRNA simultaneously.

Detailed ways

Embodiments of the present invention will be further described in detail below in conjunction with the accompanying drawings and examples. The following examples are used to illustrate the present invention, but should not be used to limit the scope of the present invention.

In this example, as shown in Figure 1-11, the preparation method of a class of functional highly branched poly(β-amino ester) disclosed by the present invention, the synthetic route is shown in Figure 1, including the following steps:

1) 4-amino-1-butanol monomer, trimethylolpropane ethoxylate triacrylate or ethylenediamine and small molecule organic amine are prepared by Michael addition reaction with double bond highly branched polymer (beta-amino ester);

2) The highly branched poly(β-amino ester) with a double bond prepared in step 1) is reacted with a functional end-capping agent to prepare a functionally highly flexible branched poly(β-amino ester).

Specifically, the following steps are included:

Add a certain amount of 4-amino-1-butanol monomer, ethylenediamine and small molecule organic amine into a two-necked glass flask with reaction solvent dimethyl sulfoxide or tetrahydrofuran and fully dissolve the monomer by magnetic stirring. The reaction molar ratio of acrylate monomer and small molecule organic amine is (1:0.5～1:3);

Further, during the reaction process, the molecular weight of the polymer is monitored by gel permeation chromatography, and the molecular weight of the polymer reaches the preset 4000Da-40000Da, and the reaction is terminated; a certain end-capping agent monomer and dimethyl sulfoxide are added to the reaction system (25°C, reaction 12h-72h), the molar ratio of capping agent and small molecule organic amine is 1:0.5~1:4, the product is purified by precipitation method, and dried in vacuum to obtain functional highly branched polymer (beta-amino ester).

Further, the diacrylate monomers used in the present invention include: 1,4-butanediol diacrylate, and the three structural formulas are as follows:

Further, the small molecule organic amine used in the present invention includes 4-amino-1-butanol, and the structural formula of 5-amino-1-pentanol is as follows:

Further, the branched monomers used in the present invention include trimethylolpropane ethoxylate triacrylate, pentaerythritol tetraacrylate, three (2-aminoethyl) amine, ethylenediamine, three [2-( Methylamino) ethyl] amine structural formula is as follows:

Further, the small molecule organic amines used in the present invention include 1-(3-aminopropyl)-4-methylpiperazine, 3,6,9-trioxaundecane-1,11- The structural formulas of diamine, 3-methylaminopropylamine and triethylenetetramine are as follows:

The functional highly branched poly(β-amino ester) prepared by the present invention is tested for the affinity performance of mRNA, and the particle size, surface potential and microscopic appearance of the formed complex nanoparticles are tested, the method is as follows:

Mix a certain amount of functional and highly flexible branched poly(β-amino ester) into the mRNA solution encoding green fluorescent protein, vortex at high speed for 20-60s, and then let it stand for 15-40min, wherein the highly branched poly(β-amino ester) The mass ratio of ester) to mRNA is 10:1-100:1, RiboGreen is used to test its mRNA affinity performance, dynamic light scattering (DLS) is used to test the particle size and surface potential of composite nanoparticles, and finally transmission electron microscopy (TEM ) to observe the microscopic morphology of composite nanoparticles.

The highly flexible branched poly(β-amino ester) of the present invention mediates the transfection efficiency and post-transfection cytotoxicity of encoding green fluorescent protein mRNA and luciferase mRNA to test, the method is as follows:

1) Cell culture: in HeLa cells, UC-3 cells, HT-29 cells, HCV-29 cells, IEC-6 cells, HepG2 cells and B16-F10 cells at a density of 1.0*104-2.0*104 cells/well Seed in 96-well plates and incubate overnight at 37°C.

2) Mix a certain amount of functional and highly flexible branched poly(β-amino ester) into the solution of coded green fluorescent protein mRNA and luciferase mRNA, vortex at high speed for 15-60s, and then let stand for 15-40min. The mass ratio of branched poly(β-amino ester) to mRNA is 10:1-100:1, and then added to the cells, and then 4-6h, the medium is replaced.

3) 24h-36h after transfection, the cells transfected with green fluorescent protein were observed and photographed under a fluorescent microscope.

4) After 24h-36h of transfection, add luciferase working solution, quickly detect the intensity of luminescence in a microplate reader, and test the transfected cell luciferase activity and the transfected cell activity.

5) Mix a certain amount of functional highly flexible branched poly(β-amino ester) into the coded green fluorescent protein DNA solution, vortex at high speed for 15-60s, and then let it stand for 15-40min, wherein the highly branched poly(β-amino ester) The mass ratio of -amino ester) to mRNA is 10:1-90:1, and then the medium mixed with complex nanoparticles containing serum is added to the cells to continue culturing for 48 hours.

Specific examples:

Example 1

The reaction ratio of 1,4-butanediol diacrylate, ethylenediamine and 4-amino-1-butanol is 3:0.2:2, and react at 90°C for 18 hours; obtain a highly branched compound with a double bond at the end Poly(beta-urethane). Then 3 times the equivalent of 1-(3-aminopropyl)-4-methylpiperazine was added and reacted at 25° C. for 48 hours to obtain functional highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da.

Fig. 1(a) is the synthesis reaction process of highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in this example. Fig. 2(a) is the H NMR spectrum of the proposed highly flexible branched poly(β-amino ester) with a molecular weight of 20,000Da prepared in this example. Figure 3(a) is the gel permeation chromatography (GPC) curve after purification of the highly flexible branched poly(β-amino ester) with a molecular weight of about 20,000Da obtained in this example, and the characteristic peaks and GPC curves presented by the proton nuclear magnetic spectrum The integrated areas of both confirmed that a highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da was successfully synthesized.

Example 2

The reaction ratio of 1,4-butanediol diacrylate, trimethylolpropane ethoxylate triacrylate and 4-amino-1-butanol is 2:0.2:2, and react at 60°C for 10 hours; To obtain a highly branched poly(β-amino ester) with a double bond at the end; then add 5 times the equivalent of 1-(3-aminopropyl)-4-methylpiperazine and react at 25°C for 48h to obtain a functional The highly flexible branched poly(β-amino ester) has a molecular weight of 15,000 Da.

Fig. 1(b) is the synthetic reaction process of the highly flexible branched poly(β-amino ester) with a molecular weight of 15,000 Da prepared in this example. Fig. 2(a) is the H NMR spectrum of the proposed highly flexible branched poly(β-amino ester) with a molecular weight of 15,000 Da prepared in this example. Fig. 3 (b) is the gel permeation chromatography (GPC) curve after purification of the highly branched poly(β-amino ester) with a molecular weight of about 15,000Da obtained in this example, and the characteristic peak position and GPC presented by the proton nuclear magnetic spectrum. The integrated areas of the curves all confirmed that a highly flexible branched poly(β-amino ester) with a molecular weight of 15,000Da was successfully synthesized.

Example 3

When the mass ratio of functional highly flexible branched poly(β-amino ester) to mRNA obtained in Example 1 was 10:1-100:1, the amount of mRNA was 0.2 μg, and the functional highly branched poly(β-amino ester) was tested by RiboGreen. Affinity properties of poly(β-amino esters) for mRNA. Firstly, the corresponding highly flexible branched poly(β-amino ester) solution is added to the mRNA solution, vortexed at high speed for 15-60s, and then left standing for 15-40min to form composite nanoparticles. Then, the nanoparticle solution was diluted to 100 μL with TE buffer solution, and then 100 μL of RiboGreen working solution (0.5%) was added, and the excited fluorescence intensity was detected under the condition that the excitation wavelength was 480 nm and the emission wavelength was 520 nm. Furthermore, a similar method was used to prepare composite nanoparticles, and the composite nanoparticles were diluted to 1mL with deionized water, and finally the particle size and surface potential of the composite nanoparticles were tested by dynamic light scattering (DLS). Freeze-drying was carried out, and the microscopic morphology was characterized by TEM.

Fig. 4 is the affinity performance test of the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000Da prepared in Example 1 to mRNA. In this example, when the mass ratio is 30:1-90:1, the implementation The highly branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in Example 1 has an affinity efficiency of more than 90% for mRNA, confirming its excellent mRNA affinity performance.

Fig. 5 is that the molecular weight prepared in embodiment 1 is the particle size test of the complex nanoparticle that the highly flexible branched poly(beta-amino ester) of 20,000Da and mRNA forms, DLS result confirms, the prepared in embodiment 1 The highly branched poly(β-amino ester) with a molecular weight of 20,000Da can effectively compress mRNA, and the particle size of the nanocomposites is less than 200nm.

Fig. 6 is the particle surface potential test of the nanocomposite formed by the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da and mRNA prepared in Example 1. DLS results confirmed that the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in Example 1 can effectively shield the negative potential of mRNA itself, and the potentials on the nanometer surface of the complex are all positive.

Fig. 7 is the microscopic morphology characterization of the composite nanoparticles formed by the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da and mRNA prepared in Example 1. The TEM results confirmed that the composite nanoparticles formed by the highly branched poly(β-amino ester) with a molecular weight of 20,000 Da and mRNA prepared in Example 1 had a uniform particle size distribution and a relatively stable structure, confirming its excellent stability.

Example 4

HeLa cells, UC-3 cells, HT-29 cells and HCV-29 cells were seeded in 96-well plates at a density of 2.0×104 cells/well, and the cells were cultured overnight at 37°C. Mix 2 μg of sodium acetate buffer solution of functional highly flexible branched poly(β-amino ester) into 0.1 μg of sodium acetate buffer solution encoding green fluorescent protein mRNA, let it stand for 45 minutes, and then slowly add it to the cells for 4 hours Then replace the culture medium. Then the cells were cultured for 32 hours, and the cells transfected with green fluorescent protein were observed under a fluorescent microscope.

Fig. 8 is the evaluation of the mRNA transfection performance of the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in Example 1. From Figure 8, it can be found that in HeLa cells, UC-3 cells, HT-29 cells and HCV-29 cells, functional highly flexible branched poly(β-amino ester) It exhibits a high GFP transfection efficiency, which proves that it can meet the high-efficiency mRNA delivery in various tissue cells.

Example 5

UC-3 cells were seeded in a 96-well plate at a density of 2.0×104 cells/well, and the cells were cultured overnight at 37°C. Mix 2 μg of functional highly flexible branched poly(β-amino ester) sodium acetate buffer solution into 0.1 μg of luciferase mRNA solution, let it stand for 45 minutes, and then slowly add it to UC-3 cells, and wait for 4 hours The medium was replaced, and the cells were cultured for 32 h. Then, the mRNA transfection efficiency was quantitatively evaluated.

For cells transfected with luciferase mRNA, wash UC-3 cells three times with PBS under dark conditions, and then add 50 μl of working solution ① (concentration: 20%) and DMEM medium mixture to UC In -3 cells, after giving for 30 min, the cell activity was detected at an excitation wavelength of 400 nm and an emission wavelength of 500 nm. Finally, 25 μl of working solution ② was added to detect the relative luminescence intensity to determine the transfection efficiency of luciferase.

Fig. 9 is an evaluation of the transfection performance of the highly flexible branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in Example 1 to mRNA encoding luciferin. The transfection results showed that in UC-3 cells, due to the advantages of multiple terminal groups, three-dimensional topology and flexible space, the functional highly flexible branched poly(β-amino ester) could efficiently mediate the expression of luciferin mRNA transfection.

Fig. 10 is the evaluation of cell viability after transfection of highly branched poly(β-amino ester) with a molecular weight of 20,000Da prepared in Example 1. From the results of cell viability, it can be found that due to highly branched poly(β-amino ester) ) good biocompatibility, degradability and extremely low dosage of highly branched poly(β-amino ester) in the process of mRNA transfection, therefore, the transfected UC-3 cells can still maintain high cell activity. This shows that functional highly branched poly(β-amino ester) has obvious advantages in mRNA delivery.

Example 6

HeLa cells, IEC-6 cells, HepG2 cells and B16-F10 cells were seeded in 96-well plates at a density of 2.0×104 cells/well, and the cells were cultured overnight at 37°C. Mix 15 μg of sodium acetate buffer solution of functional hyperbranched poly(β-amino ester) into 0.5 μg of sodium acetate buffer solution of DNA encoding green fluorescent protein, let stand for 45 minutes, and then slowly add to cells. Then the cells were cultured for 48 hours, and the cells transfected with green fluorescent protein were observed and analyzed under a fluorescent microscope.

Figure 1 is the evaluation of DNA transfection performance of highly branched poly(β-amino ester) with a molecular weight of 20,000 Da prepared in Example 1. From Figure 1, it can be found that in HeLa cells, IEC-6 cells, HepG2 cells and B16-F10 cells, functionalized highly flexible branched poly(β- Amino ester) exhibited high DNA transfection efficiency in various tissue cells, these results indicate that highly flexible branched poly(β-amino ester) can satisfy both DNA and mRNA delivery.

The embodiments of the present invention have been presented for purposes of illustration and description, but are not intended to be exhaustive or to limit the invention to the form disclosed, and although the invention has been described in detail with reference to the foregoing embodiments, it would be difficult for those skilled in the art As far as people are concerned, they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements for some of the technical features.

Claims (9)

1. A class of functional highly flexible branched polymers is characterized in that: branched polymers are β-amino esters, and the structural formula of described functional highly flexible branched polymers is as follows: In the formula, n=10-60, m=10-60; In the formula, n=10-60, m=10-60. 2. The functional highly flexible branched poly(β-amino ester) according to claim 1, wherein the molecular weight of the branched poly(β-amino ester) is in the range of 4000-40000 Da. 3. The functional highly flexible branched polyamide as claimed in claim 1, characterized in that: the molecular structure formula of the branched poly(β-amino ester) includes acrylate monomers, and the acrylate monomers The body is 1,4-butanediol diacrylate, trimethylolpropane ethoxylate triacrylate, pentaerythritol tetraacrylate. 4. The functional highly flexible branched polyamide as claimed in claim 1 is characterized in that: the molecular structural formula of the branched poly(β-amino ester) comprises small molecular organic amines, and the small molecular organic amines are 4-amino-1-butanol, 5-amino-1-pentanol 1,2-ethylenediamine, tris(2-aminoethyl)amine, tris[2-(methylamino)ethyl]amine. 5. The functional highly flexible branched polyamide as claimed in claim 1, characterized in that: the molecular structural formula of the branched poly(β-amino ester) includes an end-capping agent monomer, and the functional end-capping The agent monomer is 3,6,9-trioxaundecane-1,11-diamine, 3-methylaminopropylamine, triethylenetetramine, 1-(3-aminopropyl)-4-methyl Piperazine. 6. a preparation method applicable to the functional highly branched poly(β-amino ester) described in claim 5, is characterized in that: comprise the following steps: S1: A highly branched poly(β-amino ester) with a double bond is prepared by Michael addition reaction of a diacrylate monomer and a small molecule organic amine; S2: Prepare a functional highly branched poly(β-amino ester) by reacting the highly branched poly(β-amino ester) with a double bond prepared in step S1 with a functional end-capping agent. 7. The preparation method of functional highly branched poly(β-amino ester) as claimed in claim 6, is characterized in that: in step S1, the reaction feeding mole of described diacrylate monomer and small molecule organic amine The ratio is 1:0.5～1:3. 8. The preparation method of functional hyperbranched poly(β-amino ester) as claimed in claim 6, is characterized in that: in step S2, described functional end-capping agent reaction is that step 1) prepares with double Bonded highly branched poly(β-amino ester) and end-capping agent monomer, the total monomer concentration is 100-500mg/mL at room temperature and reacted for 12h-72h, the reaction feed mole of end-capping agent and small molecule organic amine The ratio is 1:0.5～1:4. 9. A functional highly branched poly(β-amino ester) suitable for any one of claims 1-8 is used as a cationic polymer carrier, characterized in that: the highly flexible branched functional Poly(β-amino esters) are used for mRNA delivery or DNA delivery.