Cinnamaldehyde-modified polymeric nucleic acid carrier, and preparation method and application thereof

Abstract

The application relates to the technical field of new biological medical materials, in particular to a cinnamaldehyde modified high-molecular nucleic acid carrier and a preparation method and application thereof, wherein the cinnamaldehyde modified high-molecular nucleic acid carrier is a cinnamaldehyde modified polyethylene imine polymer. The high-molecular nucleic acid carrier provided by the application can significantly improve the DNA and RNA transfection efficiency in different cells, and can effectively improve the biocompatibility of the high-molecular carrier. The preparation steps of the high-molecular nucleic acid carrier are simple, the biocompatibility is good, the repeatability of batches is strong, and the high-molecular nucleic acid carrier has a wide application prospect in the development of nucleic acid carriers, gene transfection and nucleic acid treatment and the like.

Description

Technical Field

[0001] This invention relates to the field of new biomedical materials technology, and in particular to a cinnamaldehyde-modified polymeric nucleic acid carrier, its preparation method, and its application. Background Technology

[0002] Gene therapy holds great promise for treating major diseases such as cancer. However, successful gene therapy requires highly efficient nucleic acid vectors. Polymer vectors have attracted widespread attention from scientists due to their low immunogenicity, abundant raw material sources, and modifiable chemical structures. Among them, polyethyleneimine (PEI25k), with a molecular weight of 25,000 daltons, is considered the "gold standard" of polymer vectors due to its high transfection efficiency. However, PEI25k exhibits lethal cytotoxicity, limiting its clinical application. Therefore, developing safe and efficient polymer vectors has become a key focus of gene therapy research.

[0003] Although some modification strategies can reduce the cytotoxicity of polyethyleneimine to some extent, the preparation methods are complex and have poor reproducibility. Therefore, there is an urgent need to design a safe and efficient polymeric nucleic acid vector with a simple preparation method and high reproducibility. Summary of the Invention

[0004] In view of this, the technical problem to be solved by the present invention is to provide a cinnamaldehyde-modified polymeric nucleic acid carrier, its preparation method and application, which achieves high transfection efficiency.

[0005] This invention provides a cinnamaldehyde-modified polymeric nucleic acid carrier, which is a cinnamaldehyde-modified polyethyleneimine polymer, denoted as PEI-CinA.

[0006] Specifically, the aldehyde group of cinnamaldehyde and the amino group of polyethyleneimine undergo an aldehyde-amine condensation reaction to generate an imine group, thereby achieving the grafting of cinnamaldehyde (CinA) and polyethyleneimine.

[0007] The polyethyleneimine may include one or both of linear polyethyleneimine and hyperbranched polyethyleneimine.

[0008] The molecular weight of the polyethyleneimine is preferably 300~200000 g / mol, more preferably 1000~50000 g / mol, and even more preferably 1000~30000 g / mol.

[0009] In some specific embodiments, the polyethyleneimine is selected from PEI25k.

[0010] The molar ratio of cinnamaldehyde to polyethyleneimine is preferably 1:(2~2000), more preferably 1:(2~1000), and even more preferably 1:(10~500). In some specific embodiments, the molar ratio of polyethyleneimine to carboxybenzylamine is 1:10, 1:12, 1:14, 1:20, 1:30, 1:40, 1:50, 1:100, 1:150, 1:200, 1:250, 1:300, 1:350, 1:400, 1:450, 1:500, etc.

[0011] This invention provides a method for preparing the above-mentioned cinnamaldehyde-modified polymeric nucleic acid carrier, comprising the following steps:

[0012] The cinnamaldehyde solution and polyethyleneimine solution were stirred and reacted, dialyzed, and freeze-dried to obtain a high molecular weight nucleic acid vector.

[0013] The solvent for the cinnamaldehyde solution is preferably an alcohol solvent, more preferably methanol.

[0014] In the cinnamaldehyde solution, the preferred mass-to-volume ratio of solute to solvent is 1g:(5~15)mL. For example, it can be 1g:5mL, 1g:10mL, or 1g:15mL.

[0015] The solvent for the polyethyleneimine solution is preferably one or more of water, DMSO, and alcohol solvents, and more preferably water.

[0016] The reaction temperature is preferably 15~45℃, more preferably 20~30℃, and for example, it can be 15, 20, 25, 30, 35, or 40℃.

[0017] The reaction time is preferably 6 to 24 hours, more preferably 6 to 12 hours, and for example, it can be 6, 7, 8, 9, 10, or 12 hours.

[0018] The dialysis is performed using dialysis bags well-known to those skilled in the art. Depending on the specific grafting skeleton, different molecular weight dialysis bags are used in this invention. Specifically, PEI with a molecular weight of 25000 g / mol is dialysis bag with a molecular weight of 3500 g / mol. It is preferable to change the dialysis water every 8 hours during dialysis.

[0019] The preferred dialysis time is 48-96 hours, more preferably 70-75 hours.

[0020] The freeze-drying temperature is preferably -60 to -80°C.

[0021] The present invention does not impose any special limitations on the above-mentioned vacuum drying, dialysis and freeze-drying methods, and any vacuum drying, dialysis and freeze-drying technical solutions known to those skilled in the art can be used.

[0022] In some specific embodiments, the preparation method includes the following steps:

[0023] BPEI25k was dissolved in deionized water and cinnamaldehyde was dissolved in methanol. The two solutions were then mixed and stirred to react, dialyzed, and lyophilized.

[0024] This invention provides the application of the above-mentioned cinnamaldehyde-modified polymeric nucleic acid carrier as a nucleic acid carrier in cell transfection.

[0025] The present invention also provides a polymeric nucleic acid complex nanoparticle, comprising the above-mentioned cinnamaldehyde-modified polymeric nucleic acid carrier and nucleic acid;

[0026] The nucleic acids include DNA and / or RNA.

[0027] The DNA can be double-stranded or single-stranded, and can be circular or linear, preferably pDNA. The RNA includes, but is not limited to, one or more of siRNA, mRNA, and micRNA. The RNA can be naturally isolated RNA or artificially synthesized RNA, and can be modified or unmodified RNA.

[0028] The polymeric nucleic acid complex nanoparticles can be prepared using methods well known to those skilled in the art, including but not limited to vortex recombination.

[0029] In some specific embodiments, the polymeric nucleic acid complex nanoparticles are prepared according to the following method:

[0030] The polymer-containing solution and the nucleic acid solution are mixed and vortexed to obtain the carrier / nucleic acid complex.

[0031] The concentration of the polymer in the solution is preferably 0.1~10 mg / mL, and for example, it can be 1 mg / mL.

[0032] The concentration of the nucleic acid solution is preferably 0.01~1 mg / mL, and for example, it can be 0.1 mg / mL.

[0033] The nucleic acid includes one or more of pDNA, mRNA, siRNA, etc.

[0034] The preferred volume ratio of the polymer solution to the nucleic acid solution is (1~10):1. For example, it can be 10 / 1, 5 / 1, 2.5 / 1 or 1 / 1.

[0035] The vortex duration is preferably 5 to 20 seconds, and for example, it can be 5, 10, 15, or 20 seconds.

[0036] The incubation time is preferably 5 to 20 minutes, and for example, it can be 5, 10, 15, or 20 minutes.

[0037] The aforementioned polymeric nucleic acid complex nanoparticles can be applied both in vitro and in vivo.

[0038] The preferred mass ratio of PEI-CinA to pDNA is 1~10:1; the preferred mass ratio of PEI-CinA to mRNA is 1~10:1; and the preferred mass ratio of PEI-CinA to siRNA is 1~10:1. This invention investigates the application of the vector / nucleic acid complex particles in vitro and in vivo. High transfection efficiency was achieved in various cell lines (e.g., HeLa, B16F10, MCF-7, 293T, 293F, 293S, CT26, BE(2)C, CHO, CHO-S, COS7, NIH-3T3, SKBR3, Vero, etc.).

[0039] This invention provides the application of the above-mentioned cinnamaldehyde-modified polymeric nucleic acid carrier or the above-mentioned polymeric nucleic acid complex nanoparticles in cell transfection.

[0040] Specifically, this invention provides the application of the above-mentioned polymeric nucleic acid carrier or the polymeric nucleic acid carrier or the above-mentioned polymeric nucleic acid complex nanoparticles prepared by the above-mentioned preparation method in the preparation of cell transfection reagents.

[0041] The cell transfection reagent is preferably a cationic polymer transfection reagent.

[0042] The transfection includes, but is not limited to, DNA transfection and / or RNA transfection.

[0043] The present invention also provides the application of the above-mentioned cinnamaldehyde-modified polymeric nucleic acid carrier or the above-mentioned polymeric nucleic acid complex nanoparticles in the preparation of drugs for the prevention, mitigation and / or treatment of tumors.

[0044] Based on this, the present invention also provides a pharmaceutical composition for preventing, alleviating and / or treating tumors, comprising the above-mentioned high molecular weight nucleic acid complex nanoparticles and a drug.

[0045] The drug may be an anti-tumor drug.

[0046] The tumor is preferably a solid tumor, including but not limited to melanoma, colon cancer, cervical cancer, etc.

[0047] Specifically, the exemplary steps and conditions for using PEI-CinA as a pLuc (luciferase plasmid) vector in transfection are as follows:

[0048] (1) Cell culture

[0049] Cells were cultured in a medium containing 10% fetal bovine serum and in an incubator at 37°C with 5% carbon dioxide.

[0050] (2) Cell transfection

[0051] Logarithmic growth phase cells were collected 24 hours before transfection, digested with trypsin, and diluted with 10% fetal bovine serum culture medium at a concentration of 1×10⁻⁶. 4 Cells were seeded at a density of 80%–90% in 96-well plates and incubated at 37°C in a CO2 incubator until cell confluence reached 80%–90%. For transfection, the vector / pDNA complex was incubated for 15 min, and then 0.1 μg pDNA / well was added to each 96-well plate, and the cells were cultured for another 48 h.

[0052] (3) Determination of cell transfection efficiency

[0053] Remove the cell plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer, place in -80℃ for 20 min for lysis, remove and place at room temperature, add a certain amount of luciferase substrate to each well, and quantitatively determine the cell transfection efficiency using a photometer.

[0054] (4) Detection of cytotoxicity (MTT)

[0055] The cytotoxicity of the cationic carrier / pDNA complex was evaluated using the tetramethylazozolium salt (thiazolyl blue) colorimetric method.

[0056] Logarithmic growth phase cells were collected 24 hours before transfection, digested with trypsin, and diluted with 10% fetal bovine serum culture medium at a concentration of 1×10⁻⁶. 4 Cells were seeded at a density of 80%–90% in 96-well cell culture plates and incubated at 37°C with 5% CO2 until confluence reached 80%–90%. Different mass ratios of vector / pDNA complexes were added to the cells and co-cultured for 24 h. Then, 20 μL of 0.5% thiazolyl blue solution was added to each well, and the plates were incubated at 37°C for another 4 h. The culture medium was removed, and 200 μL of DMSO was added to each well. The absorbance of the culture plates was measured using a microplate reader at a wavelength of 490 nm. Cell viability was calculated using the following formula:

[0057] Cell viability (%) = (A sample / A control )×100

[0058] A sample It is the absorption of transfected cell sample wells, A control This is the absorption of the sample pores without the addition of materials; each experiment was repeated three times.

[0059] The exemplary use of PEI-CINA as a vector for mLuc (encoding luciferase mRNA) is illustrated by the following steps and conditions:

[0060] (1) Cell culture

[0061] Cells were cultured in a medium containing 10% fetal bovine serum and in an incubator at 37°C with 5% carbon dioxide.

[0062] (2) Cell transfection

[0063] Logarithmic growth phase cells were collected 24 hours before transfection, digested with trypsin, and diluted with 10% fetal bovine serum culture medium at a concentration of 1×10⁻⁶. 4 Cells were seeded at a density of 80%–90% in 96-well plates and cultured in a constant temperature incubator at 37°C with 5% CO2 until the cell confluence reached 80%–90%. For transfection, the vector / mRNA complex was infused for 15 min, and then 0.1 μg mRNA was added to each well of the 96-well plate. The cells were then cultured for another 24 h.

[0064] (3) Determination of cell transfection efficiency

[0065] Remove the cell plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer, place in -80℃ for 20 min for lysis, remove and place at room temperature, add a certain amount of luciferase substrate to each well, and quantitatively determine the cell transfection efficiency using a photometer.

[0066] PEI-CinA, as a high-molecular-weight nucleic acid vector carrying siRNA, has been used in various cell lines that consistently express Luc (such as HeLa, B16F10, MCF-7, 293T, 293F, 293S, CT26, BE(2)C, CHO, CHO-S, COS7, NIH-3T3, SKBR3, Vero, etc.). Exemplary steps and related conditions are as follows:

[0067] (1) siRNA was prepared using conventional methods. The sequence of the siRNA was 5'-CUUACGCU GAGUACUUCGAdTdT-3', which can silence luciferase siRNA (siLuc).

[0068] (2) Cell culture

[0069] Cells were placed in a culture medium containing 10% fetal bovine serum and continuously cultured in a constant temperature incubator at 37°C and 5% CO2.

[0070] (3) Cell transfection

[0071] Logarithmic growth phase cells were collected 24 hours before transfection, digested with trypsin, and diluted with 10% fetal bovine serum culture medium at a concentration of 1×10⁻⁶. 4 Cells were seeded at a density of 80%–90% in 96-well plates and cultured in a constant temperature incubator at 37°C with 5% CO2 until cell confluence reached 80%–90%. For transfection, the vector / siRNA complex was infused for 15 min, and then 0.1 μg siRNA was added to each well of the 96-well plate. The cells were then cultured for another 48 h.

[0072] (4) Determination of cell transfection efficiency

[0073] Remove the cell plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer and lyse at -80℃ for 20 min, remove and place at room temperature, then add a certain amount of luciferase substrate to each well and quantitatively determine the cell silencing efficiency using a photometer.

[0074] The polymeric nucleic acid vector PEI-CINA provided by this invention is used for in vivo transfection and is achieved through the following scheme:

[0075] The reporter gene was selected to encode a pDNA (pLuc) that encodes luciferase. The vector / DNA complex was administered intramuscularly and intravenously to C57 mice, and the transfection effect of the complex was evaluated 48 hours after administration using in vivo fluorescence imaging.

[0076] Experimental results show that the optimal transfection efficiency of PEI-CinA carrying pDNA (pLuc) encoding luciferase in B16F10, MC38, A549, MCF-7, HeLa, and CT26 cells is 500 times that of PEI; the optimal transfection efficiency of PEI-CinA carrying mRNA (mLuc) encoding luciferase is 2000 times that of PEI; and PEI-CinA has a high gene silencing efficiency, with PEI-CinA carrying luciferase-silencing siRNA (siLuc) achieving a gene silencing efficiency of 93% in HeLa-Luc cells. Furthermore, at the optimal transfection efficiency, the survival rate in various cell types is above 95%, demonstrating excellent biocompatibility.

[0077] Compared with the prior art, the present invention provides a cinnamaldehyde-modified polymeric nucleic acid carrier, which is a cinnamaldehyde-modified polyethyleneimine polymer.

[0078] The polymeric nucleic acid vector provided by this invention can significantly improve the transfection efficiency of DNA and RNA in different cells, and can effectively enhance the biocompatibility of the polymeric vector. The preparation steps of the polymeric nucleic acid vector are simple, with good biocompatibility and strong batch-to-batch reproducibility, and it has broad application prospects in the fields of nucleic acid vector development, gene transfection, and nucleic acid therapy. Detailed Implementation

[0079] To further illustrate the present invention, a detailed description is provided below with reference to embodiments. However, it should be understood that these descriptions are merely for further illustrating the features and advantages of the present invention, and not for limiting the scope of the claims.

[0080] There are no particular restrictions on the source of any raw materials used in this invention; they can be purchased from the market or prepared using conventional methods known to those skilled in the art.

[0081] Example 1: Preparation of PEI-CinA

[0082] Weigh out BPEI25k (250 mg, 0.01 mmol) and dissolve it in 4 mL of deionized water. Dissolve a certain amount of CinA in methanol according to the designed molar ratio (see Table 1). Add the CinA solution to the BPEI solution and react at room temperature for 8 h. Dialyze the reaction mixture and freeze-dry to obtain the solid product PEI-CinA.

[0083] The grafting rates of PEI-CinA are listed in Table 1.

[0084] Table 1. Relationship between the molar ratio of raw materials and the grafting rate of CinA

[0085] Carrier material number molar ratio of PEI to CinA Number of CinA grafts PEI25k-CinA-1 1:50 43 PEI25k- CinA-2 1:100 89 PEI25k- CinA-3 1:200 187 PEI25k- CinA-4 1:300 278 PEI25k-CinA-5 1:400 346 PEI25k-CinA-6 1:500 423

[0086] Example 2: Preparation of polymeric carrier / nucleic acid complex

[0087] The initial concentration of the polymer was 1 mg / mL. The concentration of nucleic acids (pDNA, mRNA, siRNA) was 0.1 mg / mL. Volumes of the vector / nucleic acid complex were mixed according to the predetermined vector / nucleic acid complex ratios (10 / 1, 5 / 1, 2.5 / 1, and 1 / 1), vortexed for 10 seconds, and incubated at room temperature for 15 minutes to prepare the vector / nucleic acid complex.

[0088] Example 3:

[0089] (1) In vitro transfection of HeLa cells with pLuc (luciferase plasmid) mediated by PEI-CinA vector.

[0090] HeLa cell culture

[0091] HeLa cells were cultured in 10% fetal bovine serum medium and placed in a 37°C, 5% CO2 incubator.

[0092] (2) Cell transfection

[0093] Collect cells in the logarithmic growth phase. 24 hours before transfection, digest the cells with trypsin, dilute with culture medium containing 10% fetal bovine serum, and administer at a rate of 1×10⁻⁶ cells / mL. 4 Cells were seeded at a density of 80%–90% in 96-well cell culture plates and cultured in a constant temperature incubator at 37°C and 5% CO2 until cell confluence reached 80%–90%. For transfection, the vector / pDNA complex was incubated for 15 min, and then 0.1 μg pDNA / well was added to the 96-well cell plate, and the plates were cultured for another 48 h.

[0094] (3) Determination of cell transfection efficiency

[0095] Luciferase activity assay

[0096] Remove the cell culture plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer, and incubate at -80℃ for 20 min for lysis. Remove and allow to return to room temperature. Then, add a certain amount of luciferase substrate to each well, and quantitatively determine the cell transfection efficiency using a spectrophotometer. Table 2 shows the transfection efficiency of the luciferase plasmid complex.

[0097] Table 2. In vitro transfection efficiency of PEI-CinA-mediated luciferase plasmid

[0098] Carrier material number Transfection efficiency, LUC / mg Protein The mass ratio of vector to DNA PEI25k-CinA-1 <![CDATA[7.6×10 7 ]]> 2.5:1 PEI25k- CinA-2 <![CDATA[1.8×10 8 ]]> 2.5:1 PEI25k- CinA-3 <![CDATA[5.9×10 8 ]]> 2.5:1 PEI25k-CinA-4 <![CDATA[3.4×10 8 ]]> 2.5:1 PEI25k-CinA-5 <![CDATA[2.3×10 8 ]]> 2.5:1 PEI25k-CinA-6 <![CDATA[8.6×10 7 ]]> 2.5:1 PEI25k <![CDATA[2.2×10 7 ]]> 2.5:1

[0099] Example 4:

[0100] (1) HeLa cells were transfected in vitro using PEI-CinA vector-mediated mLUC (luciferase mRNA).

[0101] HeLa cells were cultured in 10% fetal bovine serum medium and placed in a 37°C, 5% CO2 incubator.

[0102] (2) Cell transfection

[0103] Collect cells in the logarithmic growth phase. 24 hours before transfection, digest the cells with trypsin, dilute with culture medium containing 10% fetal bovine serum, and administer at a rate of 1×10⁻⁶ cells / mL. 4Cells were seeded at a density of 80%–90% in 96-well cell culture plates and cultured in a constant temperature incubator at 37°C and 5% CO2 until cell confluence reached 80%–90%. For transfection, the vector / mRNA complex was incubated for 15 min, and then 0.1 μg mRNA was added to each well of the 96-well cell plate, and the plates were cultured for another 24 h.

[0104] (3) Determination of cell transfection efficiency

[0105] Remove the cell culture plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer, and incubate at -80℃ for 20 min for lysis. Remove and allow to return to room temperature. Then, add a certain amount of luciferase substrate to each well, and quantitatively determine the cell transfection efficiency using a spectrophotometer. Table 3 shows the transfection efficiency of the luciferase plasmid complex.

[0106] Table 3. In vitro transfection efficiency of PEI-CinA-mediated mLUC

[0107] Carrier material number Transfection efficiency, LUC / mg Protein The mass ratio of vector to DNA PEI25k-CinA-1 <![CDATA[1.9×10 8 ]]> 2.5:1 PEI25k- CinA-2 <![CDATA[4.3×10 8 ]]> 2.5:1 PEI25k- CinA-3 <![CDATA[7.4×10 8 ]]> 2.5:1 PEI25k- CinA-4 <![CDATA[5.7×10 8 ]]> 2.5:1 PEI25k-CinA-5 <![CDATA[2.4×10 8 ]]> 2.5:1 PEI25k-CinA-6 <![CDATA[9.2×10 7 ]]> 2.5:1 PEI25k <![CDATA[3.5×10 7 ]]> 2.5:1

[0108] Example 5:

[0109] (1) In vitro silencing experiment of HeLa-Luc cells mediated by PEI-CinA vector siLUC (siRNA for silencing luciferase).

[0110] HeLa-Luc cell culture

[0111] Cells were placed in a culture medium containing 10% fetal bovine serum and cultured continuously in an incubator at 37°C and 5% CO2.

[0112] (2) Cell silencing experiment

[0113] Logarithmic growth phase cells were collected, and 24 hours before transfection, the cells were digested with trypsin and diluted with culture medium containing 10% fetal bovine serum. Cells were then cultured at a concentration of 1×10⁻⁶ cells / mL. 4 Cells were seeded at a density of 80%–90% in 96-well cell culture plates and cultured in a constant temperature incubator at 37°C with 5% CO2 until cell confluence reached 80%–90%. For transfection, the vector / siRNA complex was infused for 15 min, and then 0.1 μg siRNA was added to each well of the 96-well cell plate, and the plates were cultured for another 48 h.

[0114] (3) Determination of cell silencing efficiency

[0115] Remove the cell culture plate from the incubator, remove the cell culture medium, wash twice with PBS, add cell lysis buffer, and incubate at -80℃ for 20 min for lysis. Remove and allow to return to room temperature. Then, add a certain amount of luciferase substrate to each well and quantitatively determine the cell transfection efficiency using a spectrophotometer. Table 4 shows the silencing efficiency of luciferase.

[0116] Table 4. In vitro transfection efficiency of PEI-CinA-mediated luciferase silencing siRNA

[0117] Carrier material number Silent efficiency Vector to siRNA mass ratio PEI25k-CinA-1 41.3% 2.5:1 PEI25k- CinA-2 78.4% 2.5:1 PEI25k- CinA-3 96.1% 2.5:1 PEI25k- CinA-4 86.2% 2.5:1 PEI25k-CinA-5 78.3% 2.5:1 PEI25k-CinA-6 68.1% 2.5:1 PEI25k 28.3% 2.5:1

[0118] (4) Assay for cytotoxicity

[0119] The cytotoxicity of the cationic carrier / pDNA complex was evaluated using the tetramethylazozolium salt (thiazolyl blue) colorimetric method. The results are shown in Table 5.

[0120] Table 5. Cytotoxicity of the PEI-CinA nucleic acid complex

[0121] Carrier material number Cell viability (%) Vector to pDNA mass ratio PEI25k-CinA-1 96.1% 2.5:1 PEI25k- CinA-2 97.3% 2.5:1 PEI25k- CinA-3 97.2% 2.5:1 PEI25k- CinA-4 96.8% 2.5:1 PEI25k-CinA-5 97.3% 2.5:1 PEI25k-CinA-6 96.7% 2.5:1 PEI25k 85.6% 2.5:1

[0122] Example 6: Application of PEI-CinA vector in in vivo pDNA transfection

[0123] (1) Cell culture

[0124] CT26 cells were placed in a culture medium containing 10% fetal bovine serum and cultured in a constant temperature incubator at 37°C and 5% CO2.

[0125] (2) In vivo transfection

[0126] Using pLuc as a reporter gene, the vector / pDNA complex was administered intravenously and intramuscularly at a dosage of 20 μg of pDNA per mouse. In vivo transfection efficiency was assessed using in vivo fluorescence imaging. Tables 6 and 7 show the in vivo transfection efficiency of the vector / DNA complex after intravenous and intramuscular administration, respectively.

[0127] Table 6. Transfection efficiency of PEI-CinA-mediated luciferase plasmid administered intravenously in vivo.

[0128] Carrier material number Transfection efficiency, LUC / mg Protein Vector to pDNA mass ratio PEI25k-CinA-1 <![CDATA[1.7×10 7 ]]> 2.5:1 PEI25k- CinA-2 <![CDATA[2.9×10 7 ]]> 2.5:1 PEI25k- CinA-3 <![CDATA[6.6×10 7 ]]> 2.5:1 PEI25k- CinA-4 <![CDATA[4.1×10 7 ]]> 2.5:1 PEI25k-CinA-5 <![CDATA[2.8×10 7 ]]> 2.5:1 PEI25k-CinA-6 <![CDATA[8.4×10 6 ]]> 2.5:1 PEI25k <![CDATA[5.2×10 5 ]]> 2.5:1

[0129] Table 7. Transfection efficiency of PEI-CinA-mediated luciferase plasmid administered intramuscularly in vivo.

[0130] Carrier material number Transfection efficiency, LUC / mg Protein Vector to pDNA mass ratio PEI25k-CinA-1 <![CDATA[1.9×10 7 ]]> 2.5:1 PEI25k- CinA-2 <![CDATA[4.8×10 7 ]]> 2.5:1 PEI25k- CinA-3 <![CDATA[7.2×10 7 ]]> 2.5:1 PEI25k- CinA-4 <![CDATA[3.5×10 7 ]]> 2.5:1 PEI25k-CinA-5 <![CDATA[2.3×10 7 ]]> 2.5:1 PEI25k-CinA-6 <![CDATA[1.4×10 7 ]]> 2.5:1 PEI25k <![CDATA[3.8×10 6 ]]> 2.5:1

[0131] The above experimental results show that the polymeric nucleic acid vector provided by this invention has high transfection efficiency in both in vivo and in vitro experiments.

[0132] The above description of the embodiments is only for the purpose of helping to understand the method and core ideas of the present invention. It should be noted that those skilled in the art can make several improvements and modifications to the present invention without departing from the principles of the present invention, and these improvements and modifications also fall within the protection scope of the claims of the present invention.

Claims

1. A cinnamaldehyde-modified polymeric nucleic acid carrier, which is a cinnamaldehyde-modified polyethyleneimine polymer.

2. The polymeric nucleic acid vector according to claim 1, characterized in that, The polyethyleneimine includes one or both of linear polyethyleneimine and hyperbranched polyethyleneimine. The molecular weight of the polyethyleneimine is 300~200000 g / mol.

3. The polymeric nucleic acid vector according to claim 1, characterized in that, The molar ratio of cinnamaldehyde to polyethyleneimine is 1:(2~2000).

4. A method for preparing a cinnamaldehyde-modified polymeric nucleic acid carrier, comprising the following steps: The cinnamaldehyde solution and polyethyleneimine solution were stirred and reacted, dialyzed, and freeze-dried to obtain a high molecular weight nucleic acid vector.

5. The preparation method according to claim 4, characterized in that, In the cinnamaldehyde solution, the mass-to-volume ratio of solute to solvent is 1 g: (5~15) mL.

6. The preparation method according to claim 4, characterized in that, The reaction temperature is 15~45℃, and the reaction time is 6~24h.

7. The preparation method according to claim 4, characterized in that, The dialysis time is 48-96 hours; The freeze-drying temperature is -60 to -80°C.

8. The use of the cinnamaldehyde-modified polymeric nucleic acid carrier according to any one of claims 1 to 3 as a nucleic acid carrier in cell transfection.

9. A polymeric nucleic acid complex nanoparticle, characterized in that, Includes the cinnamaldehyde-modified polymeric nucleic acid carrier and nucleic acid as described in any one of claims 1 to 3; The nucleic acids include DNA and / or RNA.

10. The use of the cinnamaldehyde-modified polymeric nucleic acid carrier according to any one of claims 1 to 3 or the polymeric nucleic acid complex nanoparticles according to claim 9 in the preparation of drugs for the prevention, mitigation and / or treatment of tumors.