| Chemodynamic therapy（CDT）is emerging as a therapeutic method to kill tumor cells by transforming the H2O2 in tumor cells into cells toxic hydroxyl radical（·OH）.In order to overcome the limitations of insufficient endogenous H2O2 and excessive glutathione（GSH）concentration in tumor cells,a nanoparticle Fe-MSN with CDT efficacy was designed and synthesized in this study.FHM was used to load two small molecule drugs with 3-AT and MA,respectively.Among them,3-AT serves as a catalase inhibitor to promote the accumulation of H2O2,while MA acts as a GSH scavenger to decrease the GSH content of reducing substances in tumor cells.A series of performance tests were conducted in vitro to systematically evaluate the CDT efficacy of 3-AT&MA@FHM nanoparticles.The results of 2D cell and 3D organoid experiments showed that 3-AT&MA@FHM can enhance the function of CDT and on normal cells have a certain security.The main contents of this study are as follows:（1）Synthesis of iron-doped mesoporous silica nanoparticles（Fe-MSN）.Firstly,rod-shaped iron-doped mesoporous silica（Fe-RMSN）and hollow spherical iron-doped mesoporous silica（FHM）were successfully synthesized by the one-pot method and hydrothermal methods,and the successful synthesis of iron-doped mesoporous silica was confirmed by SEM,EDS,and TEM.Meanwhile,the production of·OH was examined by electron paramagnetic resonance（EPR）.The Fe-HMSN had a good·OH production capacity at p H 6.0.In contrast,Fe-RMSN required more acidic conditions to produce·OH,which did not meet the p H requirements of the tumor microenvironment.Therefore,Fe-HMSN（hereafter referred to as FHM）was selected for subsequent experiments.（2）Study on drug loading and p H-responsive releasing of FHM.3-AT and MA were loaded with FHM,and the results of FTIR showed that 3-AT and MA were successfully loaded,and the encapsulation rates of 3-AT and MA were 62.53%±5.5%and 25.4%±1.5%by UV-vis spectroscopy.The results of the p H-responsive release of drugs in vitro showed that the maximum drug release rates of 3-AT and MA at 48 h were 24.7%and 18.7%at p H 7.4,and 60.9%and 48.0%at p H 6.0.（3）In vitro CAT inhibition,·OH production and GSH depletion studies.With the additional addition of CAT,the production of·OH was detected with 3-AT@FHM and FHM,respectively,to verify the CAT inhibition effect of 3-AT.The ability of 3-AT&MA@FHM to produce·OH by TMB colorimetric assay and the ability of GSH depletion by Ellman’s reagent（DTNB）method,respectively.The results showed that the above two effects 3-AT&MA@FHM were in line with the expected.（4）The safety and intracellular ROS production of FHM,3-AT@FHM,MA@FHM,3-AT&MA@FHM were evaluated by 2D cells experiment.The intracellular CAT inhibition ability and GSH depletion ability of 3-AT&MA@FHM were also examined.The results showed that FHM could be phagocytosed by cells and could degraded and release 3-AT and MA intracellularly,which subsequently caused intracellular CAT inhibition and GSH depletion,leading to the production and accumulation of reactive oxygen species and finally killing tumor cells,indicating that 3-AT and MA could synergistically enhance CDT with FHM.（5）A 3D organoid model evaluated the safety of FHM,and the acidic microenvironment was constructed to study the organoid-killing effect of FHM,3-AT@FHM,MA@FHM,and 3-AT&MA@FHM.The results showed that the organoid-killing effect of 3-AT&MA@FHM was similar with the trend of its cell-killing effect,in which the organoid fragmentation was caused by the gradual destruction of the cytoskeleton and a large amount of reactive oxygen species production in the cavity,which eventually led to the death of the organoid.In this study,FHM was prepared and successfully loaded with 3-AT and MA,and the ability of 3-AT&MA@FHM to enhance CDT was systematically evaluated.Meanwhile,the 3D organoid evaluation model was introduced,indicating that the 3D organoid model can be used as a rapid means of nanoparticle evaluation and has great potential in promoting the development of nanomedicine. |
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