Preparation Of Tumor Microenvironment-Responsive Nanodrugs And Their Synergistic Treatment Of Cervical Cancer

Objective:Cervical cancer is one of the most common cancers in women and the fourth leading cause of death from gynaecological malignancies.The main clinical treatments for cervical cancer are surgical resection,radiotherapy and chemotherapy,but these treatments still have shortcomings,such as high recurrence rates and systemic toxicity.The Tumor micro-environment（TME）is characterised by mild acidity,excess hydrogen peroxide（H₂O₂）,low catalase activity and high glutathione（GSH）levels,which not only provide exceptional therapeutic opportunities,but also reduce toxic effects on normal tissues.With the increasing development of nanomaterials research,the design of tumor microenvironment-responsive nanomedicines is of great importance.Therefore,in this thesis,two nanomedicines were prepared for the synergistic treatment of cervical cancer in response to the acidic environment and excess glutathione in the tumor microenvironment,respectively.Method:1.Copper sulphide（Cu S）was used as a carrier for the loading of the acoustodynamic material 5,10,15,20-tetrakis（4-aminophenyl）porphyrin（TAPP）and then coated with calcium carbonate（Ca CO₃）.The composite nanomaterial Cu S@TAPP-Ca CO₃/HA（CTCH）was synthesised by modifying the material with hyaluronic acid（HA）by electrostatic adsorption to improve the water solubility of the material.)The synthesis of CTCH was characterised using transmission electron microscope（TEM）,UV-Vis spectrophotometer,Particle size and zeta potential analyser and X-ray Photoelectron Spectroscopy（XPS）.The production of hydroxyl radicals（·OH）at different p H and H₂O₂concentrations was measured using3,3′,5,5′-tetramethylbenzidine（TMB）chromogen and methylene blue（MB）indicator,and the photothermal characterization of CTCH was carried out using an808 nm infrared laser and infrared thermography.The drug loading rate of TAPP was also measured and the calcium ion kit was used to detect the released calcium ions(Ca²⁺).In cellular experiments,the cellular uptake of CTCH was characterised using confocal microscopy,MTT,Calcein-AM/PI and other methods were used to study the cell killing effect of 2,7-dichlorohydro fluorescein diacetate（DCFH-DA）molecular probe to detect intracellular ROS production and a mitochondrial probe（JC-10）to characterise mitochondrial damage;in in vivo experiments,the anti-tumor capacity and biosafety of CTCH were assessed by body weight of mice during treatment and tumor volume,body weight,organ tissue sections,biochemical indices of mice and hemolysis assays after treatment.2.Atomic layer deposition（ALD）was used to deposit iron trioxide（Fe₂O₃）on the surface of ultrasonically treated carbon spheres（C）and polyethylene glycol（PEG）was used to coat the material to improve its biocompatibility,resulting in the synthesis of C-Fe₂O₃@PEG nanomaterials.The synthesis of C-Fe₂O₃@PEG was characterised by TEM,UV spectrophotometer,Particle size and zeta potential analyser,Inductively coupled plasma（ICP）and X-ray diffraction（XRD）.MB indicator was used to detect the production of·OH as well as to detect GSH concentration dependence,5,5’-dithiobis（2-nitrobenzoic acid）（DTNB）was used to detect the consumption of GSH by the material,and 808 nm infrared laser and infrared thermography were used to detect the photothermal properties of the material.Cellular uptake of C-200Fe₂O₃@PEG was characterised using confocal microscopy,cell killing by MTT and Calcein-AM/PI,intracellular ROS production was detected using a DCFH-DA molecular probe and intracellular GSH depletion was assessed using dialdehyde naphthalene-2,3-dicarboxaldehyde（NDA）.The anti-tumor capacity and biosafety of C-Fe₂O₃@PEG was assessed by body weight of mice and tumor volume,body weight,tumor and organ tissue sections,biochemical parameters of mice and hemolysis assays.Result:1.The successful synthesis of CTCH was demonstrated by TEM,DLS,Ze Ta potential,UV-Vis absorption,etc.The joint characterization of TMB and MB indicates that CTCH can produce·OH under acidic environment and excess H₂O₂conditions,and CTCH also has photothermal therapeutic properties;in cellular experiments CTCH has obvious ability to kill cells and also causes damage to intracellular mitochondria.CTCH has been shown to have good anti-tumor effects in in vivo experiments in mice,and there was no obvious pathological damage in tissue sections,indicating that the material is biocompatible.2.The synthesis of C-Fe₂O₃@PEG was illustrated by TEM,DLS,Ze Ta potential,UV absorption,XRD and ICP characterization.MB degradation experiments showed that C-200Fe₂O₃@PEG can produce·OH and has good photothermal properties.C-200Fe₂O₃@PEG has obvious cell The in vivo experiments in mice demonstrate that C-200Fe₂O₃@PEG has a good anti-tumor effect,and the organ tissue sections,biochemical indicators and hemolysis experiments show that the material has no obvious pathological damage to mices,indicating that the material is biocompatible.Conclusion:In this thesis,two nanodrugs were prepared that respond to the acidic environment of tumours and high GSH levels respectively.The two multifunctional nanodrugs have a selective tumour killing effect and are simple to prepare without significant toxic side effects,and have a wide range of applications in the treatment of cervical cancer and other cancers.