Development Of Targeted/Microenvironment Responsive Dual-functional LMSNs Nano-delivery System And Study On The Inhibitory Effect Against MDR Tumor

Breast cancer is one of the most prevalent cancers in the world,with 2.26 million new cases worldwide in 2020,surpassing the 2.2 million cases of lung cancer,and has replaced lung cancer as the number one cancer in the world.Surgery,radiotherapy and chemotherapy are the three traditional cancer treatments.Among them,chemotherapy is the non-specific use of chemical drugs to play a therapeutic role,the method has low targeting,high toxic side effects,low bioavailability and other disadvantages.In addition,tumors will develop drug resistance after long-term exposure to chemotherapy drugs,and this phenomenon is called multidrug resistance（MDR）,which will inevitably lead to a decrease in therapeutic index and efficacy.One of the anthraquinone antitumor antibiotics currently in clinical use,Doxorubicin Hydrochloride（DOX﹒HCl）,was selected as the chemotherapeutic agent and breast cancer cells（MCF-7）or drug-resistant breast cancer cells（MCF-7/ADR）were used as the cell lines model of this study.DOX was loaded in the pores of mesoporous silica（MSNs）.Due to the special environmental conditions such as low p H,high glutathione concentration and multiple proteins overexpression in the tumor microenvironment（TME）,we designed a disulfide bond（-SS-NH₂）on the surface of MSNs to construct a p H and reduction dual-responsive nano-drug delivery system.In order to reduce the early leakage of DOX during the transport process,liposomes were designed to coat on the surface of MSNs-SS-NH₂@DOX.In order to reverse tumor MDR and improve the targeted of the complex drug delivery system,we designed to add the modifiers vitamin E-polyethylene glycol succinate（TPGS）and hyaluronic acid（HA）on the surface of liposomes.TPGS can effectively inhibit the expression of p-glycoprotein（P-gp）on the surface of MCF-7/ADR cell membrane,and P-gp can exocytose the DOX entering the cells and produce drug resistance.HA can recognize the overexpressed CD44 receptor on the surface of tumor cells,enabling the complex drug delivery system to accumulate efficiently tumor tissues and synergistically exert anti-tumor effects.The size and micromorphology of the LMSNs composite drug delivery system were observed using particle size potentiometry,transmission electron microscope and scanning electron microscopy.The physical and chemical properties were characterized by FTIR,BET,DSC and XRD.The results showed that the prepared LMSNs core/shell composite nanocarriers have moderate particle size,uniform size and good dispersion.Functional groups such as-SS-,TPGS and HA were successfully modified into the interior or surface of the carrier.A rapid and efficient HPLC assay was established,and the results of methodology verification showed that the method could accurately quantify DOX content.The results of in vitro release study showed that the functional LMSNs drug delivery system had good p H and reduction dual response drug release characteristics.MCF-7 and MCF-7/ADR cells were studied in vitro.The cytotoxicity assay showed that the LMSNs drug delivery system could effectively kill MCF-7 and MCF-7/ADR cells.Fluorescence observation,flow cytometry,immunofluorescence staining,ROS quantitative detection,cell apoptosis and distribution cycle investigation were used to analyze and verify the efficacy of each drug delivery system against MDR tumors.The results showed that HA-TPGS-LMSNs-SS-NH₂@DOX could target tumor cells and release the drug responsively,the cell uptake rate was high,the proportion of apoptosis increased after treatment,and the growth cycle was blocked in the G0/G1 phase.The ROS generation was the highest in this treatment group.Which all indicated that HA-TPGS-LMSNs-SS-NH₂@DOX could overcome the drug resistance of tumors.MCF-7/ADR cells were inoculated into the axillae of female BALB/c nude mice,and the results of H&E staining of major organ sections showed that the LMSNs drug delivery system has high biosafety.The results of TUNEL staining and tumor inhibition rate of tumor tissue sections showed that it can effectively inhibit the growth of MDR-type tumors.After treatment with HA-TPGS-LMSNs-SS-NH₂@DOX,the tumor volume and weight of tumor-bearing mice were significantly reduced,and the survival time was significantly prolonged.The nano drug delivery system developed in this project has multiple characteristics such as targeting,microenvironment response and reversing MDR,and is expected to be applied to the efficient treatment of MDR tumors.The research results of this project can provide useful theoretical accumulation and technical support for the development of new anti-tumor nano-formulations.