Formulation And Targeting To Melanoma Cells Of Temozolomide And Multifunctional Magnetic Nanoparticles

Part ?.Formulation and targeting to melanoma cells of temozolomide loaded nanoparticlesObjective: To prepare and evaluate the temozolomide?TMZ?loaded PAMAM-based nano drug targeting delivery system,and explore it's ability of targeting to A375 melanoma cells in vitro.Methods: 1.Construction of the targeting nano delivery system of PAMAM-PEG-GE11.The PAMAM-PEG and PAMAM-PEG-GE11 were synthesized by substitution reaction and addition reaction,and the products were identified and characterized by FTIR,1HNMR,DLS and TEM,respectively.Using fluorescein isothiocyanate?FITC?modified PAMAM,and then synthesized FITC-PAMAM,FITC-PAMAM-PEG and FITC-PAMAM-PEG-GE11.Fluorescence microscopy and flow cytometry was used to check the uptake of A375 cell to the three nano materials.2.PAMAM-PEG-GE11 nano carrier for TMZ encapsulation.TMZ-PAMAM-PEGGE11-HA drug complex was prepared by “ultrasonic emulsification method”,and the particle size,Zeta potential,morphology were evaluated by DLS and TEM.In addition,the drug loading capacity and encapsulation efficiency were assayed by ultraviolet spectrophotometer.Finally,to explore if TMZ-PAMAM-PEG-GE11-HA can target to A375 melanoma cells In vitro.Results: The TMZ-PAMAM-PEG-GE11-HA targeting nano drug delivery systems was successfully synthesized by FTIR and 1HNMR.The mean particle size of it was 183.2nm and the zeta potential was-0.01 mv.The morphology was regularly spherical in shape and has good uniformity.The encapsulation efficiency of TMZ-PAMAM-PEGGE11-HA was about 50.63% and drug loading capacity of that was about 10.4%.And TMZ-PAMAM-PEG-GE11-HA can target to A375 melanoma cells in vitro research.Conclusion: The TMZ-PAMAM-PEG-GE11-HA nano delivery system has been successfully prepared,and prove its potential target to transport to the melanoma cell A375 in vitro.This drug delivery device is expected to enhance the sensitivity of melanoma to TMZ,and may provide a novel strategy for treatment of metastatic melanoma.Part ?.Multifunctional NIR Fluorescent Dyes-Magnetic Nanoparticles for Imaging and Melanoma Therapy in VitroObjective: To prepare and characterize multimodal theranostic nanoparticle,and explore it's ability of MRI and photothermal/photodynamic therapy to A375 melanoma cells in vitro.Methods: 1.Synthesis of IR820-CS-Fe₃O₄ nanoparticle:CS-Fe₃O₄ Conjugate was synthesized via high temperature coprecipitation,and then IR820 was grafted onto it to prepare IR820-CS-Fe₃O₄ nanoparticle.DLS,TEM and EDX were used to characterize them.2.Absorption spectrum determination of IR820-CS-Fe₃O₄ nanoparticle to test its stability.3.MR imaging capability of IR820-CS-Fe₃O₄ nanoparticle was measureed.4.CCK-8 assay was applied to measure the cell viability after IR820-CS-Fe₃O₄ and CS-Fe₃O₄ exposure to A375 cells;FITC-CS-Fe₃O₄ was synthesized via modification of FITC onto CS-Fe₃O₄.Flow cytometry and confocal microscopy were applied to detect the uptake and intracellular localization of nanoparticle in A375 cells.A NIR laser?808 nm?was used to irradiate the A375 cells,which were pretreated with IR820-CS-Fe₃O₄ for hours and Live-Dead Cell Staining Kit was employed to study the treatment effect.Results: We successfully prepared a theranostic nanoparticle,IR820-CS-Fe₃O₄.The average hydrodynamic size of IR820-CS-Fe₃O₄ is 24.3 nm,and its Zeta potential is 41.5 m V.The nano-material has good stability.A good T1/T2 MR imaging effect of IR820-CS-Fe₃O₄ nanoparticles was achieved in this study.CCK-8 assay showed that the nanoparticle has negligible cytotoxicity on cell viability.Flow cytometry results showed that the uptake of the nanoparticles by A375 cells in a dose dependent manner.Laser confocal microscopy results showed that the nanopartiles in cells located in lysosome.After irradiation with near infrared laser?808 nm?,the photothermal therapy and Photodynamic therapy effect was observed,resulting in a obvious killing effect on A375 cells.Conclusion: A novel multifunctional IR820-CS-Fe₃O₄ nanoparticle has been successfully prepared,and proved its excellent T1/T2 MRI capability and tumor cell killing effect,which provides a novel theranostic platform for melanoma treatment and detection.