| BackgroundCancer is one of the biggest killers threatening human life worldwide,which is a major challenge facing the current medical research field.At present,improving the effectiveness and accuracy of the tumor treatment process has become a key issue to break through the current bottleneck of tumor treatment.The development of nanoscience provides the possibility of developing new multifunctional nano-anticancer drugs.Using nanomaterials as drug carriers or directly using active nanomaterials as drugs can improve the absorption and utilization of drugs,achieve efficient target delivery,prolong the half-life of drug consumption,and reduce harmful side effects on normal tissues,thereby improving the effectiveness and accuracy of cancer treatment.Lactic acid-glycolic acid copolymer(PLGA)is a degradable functional polymer organic compound with good biocompatibility,non-toxicity,good encapsulation and film-forming properties,which is widely used in pharmaceutical and medical engineering materials And modern industrial fields.The PLGA system with different morphology,composition,structure and properties designed and synthesized at the microscopic scale through nanotechnology has been successfully applied to the diagnosis and treatment of tumors.However,PLGA itself has no drug effect,and its function is limited to use as a carrier to control the slow release of the drug contained therein and prolong the action time of the drug.How to functionalize the PLGA drug carrier is one of the effective ways to further improve the therapeutic effect of the drug loading system.Nanozyme is a kind of nanomaterial with inherent enzyme-like activity,whose multi-enzyme activity can regulate the level of reactive oxygen species(ROS)in biological systems.For example,superoxide dismutase(SOD)and catalase(CAT)activities can be used to remove ROS,while peroxidase(POD)and oxidase(OXD)activities can be used to generate ROS.It is worth emphasizing that nanozymes with peroxidase and oxidase activities can catalyze the oxygen and hydrogen peroxide in tumors to produce highly toxic reactive oxygen radicals to achieve specific killing of tumor cells.If the nanozyme and the PLGA drug carrier are combined,it is expected to obtain a functionalized PLGA nanocarrier,which can release the drug inside the carrier for tumor chemotherapy as well as exert the effect of nanozyme catalyzed tumor therapy.The two effects are synergistic with each other,thereby improving the effectiveness of cancer treatment.Based on the above research background,this paper first functionally modified PLGA nanoparticles,modified the manganese oxide compound(MnOx)with oxidase activity to the surface of PLGA nanoparticles,thus preparing PLGA-MnOx composite nanoparticles,and evaluating the morphology,particle size,structure,multi-enzyme activity,and the ability to catalyze the killing of tumor cells in detail.Subsequently,the anti-tumor drug Artesunate(ARS)was loaded inside the PLGA-MnOx composite nanoparticles to investigate the inhibition of tumor cells by the synergy of chemotherapy and catalytic therapy.Finally,the animal model was used to evaluate the tumor treatment effect.PurposeThis article takes nanosynthetic chemistry as a means and uses the application in the medical field as a guide to functionally modify PLGA nanopharmaceutical carriers and modify MnOx nanozymes with multiple enzyme activities to the surface of PLGA nanopharmaceutical carriers,thereby constructing tumor catalytic Nano-drugs with synergistic effects of chemotherapeutics to improve the effectiveness of cancer treatment.MethodsChapter 1 Preparation and Multi-enzyme Activity of PLGA-MnOx Nanozyme In this chapter,using PLGA as the nucleating material,the PLGA nanoparticles are prepared by the W/O/W emulsification method,whose physical and chemical properties are analyzed by various detection methods,including size distribution and surface properties.During the experiment,the composition,size and morphology of PLGA-MnOx nanozymes were adjusted by controlling the input amount of potassium permanganate and the reaction temperature.The morphology of PLGA-MnOx nanozyme was characterized in detail by means of transmission electron microscope(TEM)and scanning electron microscope(SEM).Subsequently,the structure and composition of the material were analyzed by X-ray diffraction spectroscopy(XRD)and X-ray photoelectron spectroscopy(XPS).Finally,the multi-enzyme activities of PLGA-MnOx nanozymes were analyzed,including oxidase,catalase,peroxidase,superoxide dismutase and so on.Chapter 2 ARS@PLGA-MnOx Nanozyme for tumor catalysis/chemotherapyIn this chapter,the artesunate package is loaded into the PLGA-MnOx nanozyme.The anti-tumor activity of drug-loaded nanozymes in vitro was evaluated by MTT and live cell staining methods,including the killing ability of ARS@PLGA-MnOx nanozymes on tumor cells,the uptake experiment of PLGA-MnOx nanozymes by tumor cells,intracellular ROS detection of levels and mitochondrial membrane potential,evaluation of in vivo and in vitro MRI effects,etc.Finally,subcutaneously transplanted tumor models of mouse colon cancer(CT26)and mouse breast cancer(4T1)were constructed to evaluate the effects in vivo.ResultChapter 1:The PLGA-MnOx diagnostic nanozyme prepared in this chapter is a sphere with uniform size,a particle size of about 180 nm with good dispersion.Through XPS analysis,it was found that PLGA-MnOx nanozyme has the existence of various elements such as C,N,O,Mn,which proved the successful incorporation of Mn in the structure.In addition to the three enzyme activities common to nanomaterials:oxidase,catalase,and superoxide dismutase,PLGA-MnOx nanozymes also have the ability to remove glutathione.The results of Mie equation analysis showed that compared with the control of PLGA and PLGA-PDA,various enzyme activities of PLGA-MnOx were significantly increased,indicating that Mn doping wasthe entire source of PLGA-MnOx nanozyme enzyme activity.Chapter 2:This chapter combines artesunate with PLGA-MnOx nanozyme to prepare a drug-loaded composite nanozyme(ARS@PLGA-MnOx)with a drug loading rate of 13.9%.In addition,MTT results also showed that the survival rate of tumor cells decreased with the increase of ARS@PLGA-MnOx concentration,which proved that the combination of artesunate and PLGA-MnOx had a synergistic inhibitory effect on tumors.At the cellular level,DCFH-DA fluorescent probe experiment results show that PLGA-MnOx nanozymes can produce more reactive oxygen species in cells than blank PLGA nanoparticles.The mitochondrial membrane potential test results and Western Blot results can show that PLGA-MnOx and ARS@PLGA-MnOx can induce more tumor cell apoptosis.In CT26 tumor-bearing mice and 4T1 tumor-bearing mouse models,ARS@PLGA-MnOx can inhibit tumor growth more than PLGA-MnOx.The mouse survival rate experiment also showed that the mice treated with ARS@PLGA-MnOx showed higher survival rates than the two groups of mice in the PLGA-MnOx alone administration group and the ARS alone administration group.The results of histochemical analysis showed that PLGA-MnOx and ARS@PLGA-MnOx did not cause signi-ficant damage to the main organs of mice,indicating that they had no obvious toxic and side effects. |
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