Targeting Phase-transition Nanoparticles For Multimodal Imaging And Sonodynamic-gene Therapy For Thyroid Papillary Carcinoma

Background: Papillary thyroid carcinoma(PTC)has a relatively high incidence rate,and some tumors have the risk of lymph node metastasis at an early stage.At present,the common methods of diagnosis and treatment of PTC have their own advantages and disadvantages,and there is controversy about excessive diagnosis and treatment.In order to achieve accurate diagnosis and treatment of PTC,a phase-transition nanoparticles(PFP@IR780/PLGA-b PEI-miRNA338-3p)was designed based on perfluorocabon nanoparticles(PFC NPs).The nanoparticles can target the tumor cells,realize the multimodal imaging,and improve the accuracy of tumor diagnosis.Moreover,the nanoparticles can also target transfection of miRNA338-3p,and generate cytotoxic reactive oxygen(ROS)under the effect of low intensity focused ultrasound(Li FU),which can act on tumor cells in combination to realize sonodynamic-gene therapy for PTC,so as to achieve the purpose of precise diagnosis and treatment.Objective: A cationic polymer compound PLGA-b PEI was used as a spherical shell,and the shell was loaded with IR780.The core of the shell was Perfluoropentane(PFP),and the surface was bonded with miRNA338-3p to prepare the targeted phase transformation nanoparticle P@IP-miRNA.The basic performance of PTC was tested,and the multimodal imaging ability of nanoparticles in vitro and in vivo was evaluated,and the sonodynamic-gene therapy effect of nanoparticles in vitro and in vivo was studied.Methods: Nanoparticle was prepared by double emulsification method,and miRNA338-3p was attached to the surface of the nanoparticles by electrostatic adsorption method.The morphology,size and dispersion of nanoparticles were observed by inverted fluorescence microscope,scanning electron microscope and transmission electron microscope.The particle size and potential of the nanoparticles were measured by Malvin particle size analyzer.The encapsulation rate,drug loading rate and ROS production ability of IR780 after irradiation of LIFU were measured by ultraviolet spectrophotometer.The connection between the nanoparticles and miRNAs was observed under a confocal microscope.Fluorescence labeling method,flow cytometry and micro UV spectrophotometer were used to detect the adsorption efficiency of miRNA nanoparticles.In vitro experiments,laser confocal microscopy and flow cytometry were used to detect the in vitro targeting and subcellular localization of nanoparticles,and q RT-PCR,Western blot and immunofluorescence experiments were used to detect the miRNA transfection ability of nanoparticles.CCK8 kit was used to evaluate the safe concentration and inhibitory effect of nanoparticles on TPC-1 cells.Laser confocal microscopy and flow cytometry were used to detect the apoptosis of TPC-1 cells under different treatments,and to evaluate the efficacy of combined therapy in vitro.The multimodal imaging ability of nanoparticles in vivo and in vivo was detected by ultrasonic diagnostic instrument,fluorescence imager of small animals and photoacoustic imager,and the distribution of nanoparticles in vivo,tumor targeting and the optimal time window of combination therapy were analyzed.In vivo treatment,in vivo biosafety of nanoparticles was detected.The body weight and tumor size of the tumor-bearing nude mice were monitored and recorded under different treatment groups.The tumor inhibition rate of each group was calculated,and pathological detection was performed to comprehensively evaluate the efficacy of combined treatment with nanoparticles in vivo.Results: P@IP-miRNA(PFP@IR780/PLGA-b PEI-miRNA-338-3p)nanoparticles were successfully prepared with spherical shape,uniform size,good dispersion and positive potential.The encapsulation rate of IR780 was(82.58±3.92)%,the drug loading rate was(6.60±0.32)%,and the adsorption capacity of miRNA338-3p was 41.78 ?g/mg.Nanoparticle has excellent tumor targeting ability,miRNA transfection ability,ROS production ability and multimodal imaging ability in vivo and in vitro.In vivo studies,the biosafety of P@IP-miRNA was good.The antitumor effect of P@IP-miRNA+LIFU combined treatment group was the best,and the efficacy was better than that of single factor treatment group,and the difference was statistically significant.Pathological detection and immunohistochemical analysis of tumor tissues and major organs in each group were conducted,and the results were consistent with the treatment results.Conclusion: In this study,the targeting phase-transition nanoparticles(P@IP-miRNA)was successfully prepared.The nanoparticle has good targeting ability to tumor and has biological safety,which can realize multimodal imaging and sonodynamic-gene therapy for PTC,providing a new idea for accurate diagnosis and treatment of PTC.