Construction And Therapeutic Performance Evaluation In Vitro Of Multifunctional Nanotherapeutic System For Enhanced Chemodynamic Therapy

Cancer,also known as malignant tumor,is a disease in which normal cells proliferate and differentiate abnormally and develop into tumor tissue.It has become one of the main causes of human death.The strategy of clinical treatment of cancer is to remove tumor lesions and kill cancer cells.The main methods include surgery,radiotherapy and chemotherapy.However,these methods have some defects,such as great trauma,low selectivity or insufficient curative effect.In recent years,new therapies such as photodynamic,photothermal and chemodynamic therapy have been proposed,which have good therapeutic effect,strong specificity and less trauma,and have a prospect of clinical application.However,due to the complexity of tumor microenvironment and the limitations of these new treatment methods,a single treatment method is still difficult to effectively kill cancer cells and inhibit tumor growth.The combined use of different therapies to kill cancer cells can not only overcome the shortcomings of single treatment,but also weaken the side effects of treatment and greatly enhance the therapeutic effect.As a new therapeutic material,nanomaterials have controllable size and morphology,adjustable optical properties and easy multifunctional modification.They play an important role in the research of photodynamic,photothermal,chemodynamic and their synergistic therapy.Based on the unique properties of silica,metal organic framework and upconversion nanomaterials,aiming at the problems of limited chemodynamic treatment caused by tumor microenvironment and poor treatment effect of single treatment mode,and in order to synergistically enhance chemodynamic treatment,the nanosystems of chemodynamic/photodynamic therapy,chemodynamic/chemotherapy and chemodynamic/photothermal/chemotherapy were designed and prepared to achieve synergistic and efficient killing of cancer cells.The research contents are as follows:（1）In view of the low efficiency of Fenton reaction for chemodynamic therapy in tumor microenvironment and the restriction of hypoxia on photodynamic therapy,a photo-chemodynamic/photodynamic combined therapy nanomaterial（UCTSCF）was designed by using the near-infrared light（808 nm）driven upconversion nanomaterial（UC）as the matrix,the photosensitizer chlorin e6（Ce6）/oxygen carrying triethoxy（1H,1H,2H,2H-nonafluorohexyl）silane（TFS）and copper doped iron-based MOFs（MIL-100（Cu/Fe））as tandemly modified functional layers.Under 808 nm light excitation with high tissue penetration,UC emits strong blue fluorescence and activates MIL-100（Cu/Fe）reacting with hydrogen peroxide（H₂O₂）in the tumor microenvironment to produce·OH,realizing efficient the effect of photocatalytic chemodynamic（photo-chemodynamic）therapy.This result was confirmed by methylene blue（MB）degradation experiment.The photosensitizer Ce6 contained in UCTSCF is rapidly activated under 671 nm laser irradiation,and the oxygen carrier TFS provides sufficient O₂ to generate a large amount of singlet oxygen（¹O₂）to realize the photodynamic treatment of oxygenation.1,3-diphenylisobenzofuran（DPBF）was used as a probe to verify the large amount of ¹O₂produced.UCTSCF enters cancer cells through endocytosis.Due to photo-chemodynamic/photodynamic combination therapy,the lethality rate of breast cancer cells（MCF-7）reaches 80%at 24 hours,much higher than 48.7%of single photo-chemodynamic therapy and 61.5%of single photodynamic therapy.This indicates that UCTSCF has great potential for efficient photo-chemodynamic/oxygen-elevated photodynamic therapy.（2）In order to improve the catalytic activity of Fenton reagent in tumor microenvironment,hypoxia and weak penetration of ultraviolet/visible light in photodynamic therapy,a combined chemodynamic/photodynamic therapeutic nanomaterial（UCTSH）was constructed by using an intense near-infrared light（808 nm）driven red-emitting upconversion nanomaterial（UC）as the matrix,the photosensitizer Ce6/oxygen carrying-TFS and copper based MOFs（HKUST-1）as tandemly modified functional layers.Under weakly acidic conditions,copper based MOFs in the outer shell of UCTSH degrade and release Cu²⁺,which catalyzes the endogenous H₂O₂ of tumor cells to produce·OH,and achieve the effect of chemodynamic therapy.Besides,under the irradiation of 808 nm laser,UC emits red light to activate photosensitizer Ce6,which can convert the endogenous and TFS released oxygen to¹O₂,leading to highly efficient photodynamic therapy.DPBF degradation experiment was used to verify the synergistic production of·OH and ¹O₂ by UCTSH.In addition to the oxygen carrying capacity,TFS can also protect the upconversion luminescence of UCTSH.Compared with the single photodynamic or chemodynamic therapeutic nanoagent in the literatures,the UCTSH shows higher mortality to MCF-7 cells under smaller therapeutic dosage（24-hour survival rate is only 14.1%）,indicating that this material is an excellent nanoagent combined with high-efficiency chemodynamic and oxygenation photodynamic therapy.（3）Aiming at the problems of low H₂O₂ content in tumor microenvironment,insufficient·OH generated and uncontrollable drug release in chemotherapy,a smart p H triggered combined chemodynamic/chemotherapy treatment nanosystem（DMSNs-DOX-CP）was designed by co-loading copper peroxide（CP）nanodots and chemotherapeutic drug DOX onto dendritic mesoporous nanosilica（DMSNs）.Due to its large pore size and specific surface area,the load of DMSNs on CP and DOX is as high as 80.0%and 19.8%respectively.Under weak acidic conditions,DMSNs-DOX-CP can release a large amount of Cu²⁺and H₂O₂,addressing the deficiency issue of endogenous H₂O₂ in tumors to produce more·OH through Fenton-like autocatalytic reaction,leading to enhanced chemodynamic therapy.Moreover,DOX release of DMSNs-DOX-CP increased significantly in the simulated tumor micro acid environment,suggesting a potential targeted therapeutic effect.The 24-hour survival rate of cervical cancer cells（He La）was as low as 15.3%in the presence of low dosage of DMSNs-DOX-CP（100μg/m L）,which was much lower than that of the single chemodynamic treatment（44.7%）and single chemotherapy treatment（52.1%）,indicating efficient synergistic therapy of the as-designed nanotherapeutic system.（4）In order to solve the problem of low efficiency of single chemodynamic therapy,Fe²⁺doped ZIF-8 ultra-small nanocrystalline carrier（FZ）was prepared by ion doping strategy,and then a nanotherapeutic system（FZID）with synergistic chemodynamic/photothermal/chemotherapy was constructed by co-loading photothermal agent indocyanine green（ICG）and chemotherapeutic drug DOX in one pot.The degradation of ZIF-8 in acidic environment render quick Fe²⁺release（30 min,the release amount is 82.7%）of the FZID.The released Fe²⁺can catalyze H₂O₂ to produce a large amount of·OH（Fenton reaction）per unit time.Compared with other similar materials,the FZID shows a higher Fenton reaction rate and improves the effect of chemodynamic treatment.On the other hand,the ICG loaded in FZID can absorb the excitation light of 808 nm to elevate the temperature with a photothermal conversion efficiency of 35.9%,which is much higher than free ICG（15.8%）and some widely used photothermal therapeutic agents in the literatures.It shows that FZID has high photothermal treatment efficiency.At the same time,the hyperthermia effect of FZID improves the Fenton reaction rate and realizes the photothermal effect-enhanced chemodynamic therapeutic.Additionally,the FZID can release DOX（20 min,81.1%）rapidly to supply efficient chemotherapy.The 24-hour lethality rate of this photothermal/chemotherapy/enhanced chemodynamic nanosystem to He La cells is as high as93.7%,which is much higher than any of the single treatment mode.The nanosystem also demonstrates little damage to normal cells（the survival rate is 75.0%）.Results indicate the FZID has good antitumor therapeutic effect in vitro and is expected to be applied to antitumor research in vivo.