Preparation And Characterizaion Of Acid-Sensitive Prodrugs Based On Polyethylene Glycol And Modified Dextran For Co-Delivery Of Doxorubicin And P53 Gene

Cancer is a major disease that threatens human health,and the traditional treatment methods are not enough to inhibit the development of tumors.The rapid development of nanotechnology in the development of nanomedical products has brought great potential for improving cancer treatment strategies,which can improve the pharmacokinetic and pharmacodynamic characteristics of conventional therapeutic agents,and thus optimize existing anticancer drugs.The nanocarrier has the advantages of nanometer scale,high specific surface and good physical and chemical properties.Using nanocarriers to transport drugs can improve the stability of drugs in vivo,prolong the blood circulation time,and achieve the controlled release of drugs.In addition,nanocarriers can alter the biological distribution of drugs and enrich drugs at tumor sites by the enhanced permeability and retention(EPR)effect.However,due to the complexity of cancer pathology,a single type of treatment cannot effectively cure cancer fundamentally.Therefore,combined therapy as a new anti-tumor treatment method has been widely studied.A combination of genes and drugs often produces better results than a single drug treatment,and reduces drug dependence and pain.Tumor suppressor gene p53 can induce apoptosis or cell senescence,and plays a crucial role in inhibiting cell proliferation,so it has been widely studied as a therapeutic gene.1.Preparation and characterization of acid-sensitive doxorubicin prodrug based on polyethylene glycol.In order to solve the problem of cumbersome preparation drug carriers and low drug loading,we designed and prepared a PEG-based pH-sensitive polymeric prodrug DOX-PEG-DOX,which has simple structure and easy to prepare via Schiff-base reaction.Firstly,aldehyde group-modified ALD-PEG-ALD was carried out by esterification reaction using polyethylene glycol(PEG)with molecular weights of 1 k,2 k and 4 k,respectively.And then acid-sensitive prodrug DOX-PEG-DOX were obtained by a Schiff-base reaction.The structure of prodrugs were characterized by magnetic resonance spectroscopy(1H NMR),fourier transform infrared spectroscopy(FT-IR)and high performance liquid chromatography(HPLC).The amphiphilic prodrugs were able to self-assemble into nanoparticles in water.The particle size and morphology of the nanoparticles were observed by dynamic light scattering(DLS)and transmission electron microscopy(TEM).The stability of nanoparticles at pH 7.4 and the particle size change at pH 5.0 were tracked by DLS.The drug release behavior under different pH conditions was studied by in vitro drug release experiment.In order to test the inhibitory effect of prodrug on tumor cells,different doses of prodrug were added to HeLa and HepG2 tumor cells to evaluate the inhibitory effect on tumor cells.Meanwhile,the endocytosis of prodrug nanoparticles into cells and the process of drug release were observed intuitively by endocytosis experiments.The above studies have proved that this kind of pegylated prodrugs with simple structure have has promising potential in the application for cancer treatment.2.Functional dextran for co-loading of DOX and p53 gene.Dextran is a natural polysaccharide with similar properties to PEG,and it also has broad application prospects in biomedical materials.First,the dextran was modified with alkynyl group,and then the hydrophobic polycaprolactone(PCL)and the protonatable tertiary amine group were grafted onto the dextran by a two-step click reaction to construct a cationic polymer carrier DAE-Dex-g-PCL that can co-load DOX and p53 genes.The structure of DAE-Dex-g-PCL was characteriz ed by 1H NMR.The cationized dextran could self-assemble in water and simultaneously package DOX,and then the p53 gene was compressed by electrostatic action to form composite nanoparticles(D/P-NPs).We used DLS to observe the particle size and measured the zeta potential under different N/P ratios.The assembly morphology of the composite nanoparticles was observed by TEM.The ability of immobilized p53 gene and the stability of nanoparticles were tested by gel retardation electrophoresis.In vitro drug release experiment was carried out to study the drug release rate.Finally,the inhibition effect of the composite nanoparticles on tumor cells(H1299 cells)was studied by cytotoxicity test,and the release process of drug and p53 gene in H1299 cells was observed intuitively by endocytosis experiments.The above results indicate that the modified dextran has an important application prospect in the study of anti-tumor.