Study Of Multiple Pathways Synergistic Interference With Drug-resistant Tumor Cell Microenvironment To Enhance The Efficacy Of Doxorubicin

Malignant tumor is a severe disease threatening human health.Chemotherapy is the main means to treat malignant tumor at present.However,drug resistance associated with chemotherapy is one of the main reasons giving rise to the failure of cancer treatment.In order to overcome tumor drug resistance,many treatment methods have been studied,such as reducing the drug efflux by inhibiting P-glycoprotein（P-gp）and disturbing the redox balance in tumor cells by reducing the level of glutathione（GSH）.However,due to the complex microenvironment in tumor cells,a single interfering method is difficult to obtain ideal therapeutic effect.Therefore,it is urgent to develop new treatment methods to solve the problem of drug resistance by interfering tumor microenvironment in the process of chemotherapy,and improve the efficacy of antitumor drugs.Since tumor drug resistance is closely related to the microenvironment of tumor cells,interfering the microenvironment of tumor drug-resistant cells through multiple pathways at the same time may reduce the tolerance of drug-resistant tumor cells to chemotherapeutic drugs.Therefore,a drug delivery system was designed that GSH responsive metal organic framework（MOF）co-deliver doxorubicin（Dox）and photosensitizer modified antisense oligonucleotides of HIF-1α.This nanosystem can consume GSH in drug-resistant tumor cells,thus interfere the intracellular redox balance and inducing ferroptosis to resist drug resistance.Antisense oligonucleotides were also used to inhibit the expression of P-gp through gene therapy and reduce the efflux of chemotherapeutic drugs.Moreover,the loaded photosensitizer can be used for photodynamic therapy（PDT）,which interfere the mechanism of cell apoptosis and promote cell apoptosis.Therefore,this strategy of multiple pathways can be applied to interfere the microenvironment of drug-resistant cancer cells and improve the therapeutic effect of Dox on drug-resistant breast cancer.The main contents and experimental results of this study are as follows:firstly,new MOF nanoparticles containing disulfide bonds were synthesized and characterized by TEM,BET and XRD.And the concentration range of rational use of the material was explored,the loading amount of antisense oligonucleotide was 200 n M.After that,the constructed MOF nanosystem was characterized.The drug loading capacity was 15.2%and the drug release capacity was 78%.The inhibition of tumor cells by the nanosystem is determined by the MTT assay.The experimental results showed that the inhibition rate of tumor cells reached 84%after combined PDT treatment when MOF（200μg/m L）loaded with Dox（5μg/m L）and photosensitizer modified antisense oligonucleotide（200 n M）.At the same time,the IC₅₀of Dox to MCF-7/ADM is 0.87μg/m L under this nanosystem,much less than the IC₅₀of free Dox to MCF-7/ADM（128.3μg/m L）.Moreover,the cell killing effect of nanosystem was further verified by cell apoptosis detection kit and live and dead cell detection kit.Then,the uptake ability of tumor cells to MOF delivery system was studied by laser confocal experiment,and the results showed that tumor cells could rapidly uptake the nanosystem after incubating with the nanosystem for 6 hours.Western blot showed that the nanosystem could downregulate the anti-apoptotic protein（Bcl-2）,hypoxia inducible factor-1（HIF-1α）,P-glycoprotein（P-gp）and glutathione peroxidase（GPx4）,the expression was dose-dependent with the concentration of antisense oligonucleotides.Reactive oxygen species（ROS）detection kit was used to explore the effect of nanosystem on inducing ROS in tumor cells.The GSH detection kit proved that such MOF nanoparticles had an excellent ability to consume the content of GSH in cells,which was dependent on the addition time of MOF.At the same time,the mouse tumor model in vivo was constructed to evaluate the antitumor activity of the nanosystem in vivo.The distribution of drugs in mice under different treatment methods was detected by near infrared fluorescence optical imaging system,which verify the tumor targeting ability of nanosystem in vivo.Finally,H&E staining was used to evaluate the systemic toxicity of the nanosystem through histopathology.To sum up,the multifunctional nanosystem constructed in this study interferes the microenvironment of tumor cells through multiple pathways,which enhances the drug sensitivity of drug-resistant tumor cells to chemotherapeutic drug.This strategy shows excellent effect of anti drug-resistance,and provides a novel strategy for more efficient treatment of drug-resistant tumors.