Design And Biological Applications Of Photoactive Polymer-Nucleic Acid Nanocomposites

Singlet oxygen(1O2),as a special kind of reactive oxygen species(ROS)with higher oxidation activity,has been widely studied and applied in biomedical fields such as photodynamic therapy(PDT)and photochemical internalization(PCI)-mediated gene delivery.In this paper,we first utilize spiropyran-decorated polymers to construct a photosensitive nanosystem with reversible photosensitizing capability and further investigate its potential applications in PDT.Secondly,a small amount of spiropyran-containing polymer photosensitizer was applied to non-covalently modify the polymeric shell of the bPEI/DNA polyplexes by sequential self-assembly method with the hope of solving the problems of high toxicity of bPEI and low transfection efficiency under low DNA dosage conditions using PCI effect and effectively avoiding the non-specific photochemical toxicity of traditional photosensitizers.In addition,the regioregular cationic polythiophenes were employed to construct a photosensitive gene delivery system and its ability to enhance gene delivery was further investigated.The paper mainly includes the following three parts:Part 1.Reversibly controlled generation of singlet oxygen from photosensitizing nanosystems is a challenging option for photodynamic therapies.For the first time,spiropyran units were used with imaging and oxygen photosensitization properties in nanoassemblies formed by spiropyran-containing cationic copolymers and plasmid DNA.By virtue of photochromic property of spiropyran units,the nanoparticles indicated multiple merits on the side of spiropyran chromophore:1)a suitable MC half life of up to 2.8 h in the dark;2)relatively high Фf(with a quantum yield of up to 0.27);3)applicable Ф△(with a quantum yield of up to 0.22);4)reversible control in fluorescence and ROS generation by light,and 5)dual role in cell imaging and ROS-induced cell apoptosis.We found spiropyran-containing nanoparticles had photoswitching properties in both fluorescence and singlet oxygen generation in aqueous solutions and cells,and demonstrated reversible aggregation-induced enhanced photosentization and emission could be potentially applied for photodynamic therapy studies.Part 2.PCI is a technology that allows light-induced delivery of DNA,drugs or other biological factors directly inside cells.In this study,a novel PCI-mediated gene delivery system was developed through the sequential self-assembly of plasmid DNA(pDNA),branched polyethylenimine(bPEI),and photosensitizer(PS),forming polyplex with three-layered functional nanocompartments.This is a ternary polyplex composed of a core containing pDNA packaged with bPEI and spiropyran-containing cationic copolymers P2 or P3,which anchored at the outermost layer and provide the photosensitizing action as PS with capability of reversibly control generation of 1O2 and featured non-photosensitive upon natural light irradiation after PCI treatment.Because the PS and DNA cargoes were compartmentalized distantly in the polyplex shell and core,respectively,the generated 1O2 caused minimal damage to DNA molecules to preserve their transfection potency.Under almost non-toxic ternary polyplexes-mediated PCI treatment,the expression efficiency of green fluorescent protein(GFP)with lower DNA usage determined by flow cytometry was markedly enhanced from 15.4%to 91.2%in HeLa cells.Therefore,the ternary polyplexes with suited irradiation is a promising non-toxic and photo-inducible effective gene delivery strategy,which should be encouraged in tumor therapy.Part 3.Polythiophene as a class of conjugated polymers with excellent photosensitizing activity attracts much attention in the biomedical field.We use regioregular cationic polythiophene as a polymer photosensitizer and DNA condensation agent to explore its application prospects in the field of PCI-mediated gene delivery.Compared to previously reported nonregioregular polythiophenes,S1-S4 exhibited enhanced DNA binding and condensing capability.The gene delivery performance of S1-S4 was significantly improved by introducing PCI effect with polythiophene backbones as photosensitizer to overcome endolysosome barrier while maintaining integrity of complexed DNA,which may alleviate the concern of ROS-assisted transgene enhancement by shortly elevating ROS level in endolysosome compartments.