A Preliminary Research On Molecular Label Recognition-triggered Drug Delivery System

Malignant tumor is one of the important threats to people's life and health in China.Chemotherapy is the main clinical treatment for malignant tumor.However,chemotherapy drugs are limited by the biological barrier in vivo,toxic side effects,and drug resistance.In recent years,the intelligent drug delivery system of tumor has become a research hotspot.To a certain extent,it improves the behavior and distribution of chemotherapy drugs in vivo and improves the therapeutic effect.However,the current reported intelligent drug delivery system based on tumor microenvironment?pH,GSH?or micro-energy?near-infrared,ultrasound?is mostly limited by tumor types,tumor locations and degree of tumor development,resulting in limited response degree and drug release to achieve efficient killing of tumor cells.Based on the above problems,it is urgent to strategically design and construct a tumor cell molecular tag recognition-trigger intelligent drug delivery system.This project intends to construct and prepare a kind of mesoporous nanomaterial,in which the surface is modified with ligand,which has the ability of molecular recognition and spatial conformational change.The molecular recognition of tumor cells can occur simultaneously with controlled drug release,so as to trigger drug release and activate chemotherapy.The drug delivery system was composed of a drug storage unit and an intelligent identification and responsive drug release unit.Mesoporous Silica Nanoparticles?MSNs?were used as a drug storage unit because of its high biological safety,simple preparation process,adjustable pore size,extra large specific surface area,easy surface modification and ultra-high drug loading.Aptamer?APT?was used as an intelligent identification and response release unit based on its target specific affinity,easy molecular engineering transformation and conformational predictability.The hairpin APT with remarkable molecular recognition and conformational change ability was designed and prepared with the specific marker Mucin 1?MUC-1?on the surface of breast cancer cells.An intelligent drug delivery system based on molecular tag recognition and trigger was constructed.MSNs were synthesized by sol-gel method,then azide was modified on the surfaces of MSNs?MSNs-N₃?,after which the antitumor drug Doxorubicin?DOX?was load into MSNs-N₃.Finally,with the click reaction of alkynyl at the tail of hairpin APT reacting with azide on the surfaces of MSNs,specific recognition of tumor cell surface label protein MUC-1 was modified with hairpin APT.On the one hand,the hairpin APT served as a"gate control"to avoid DOX leakage;on the other hand,the hairpin APT could specifically recognize MUC-1 protein.More importantly,it could induce conformational changes in hairpin structure at the same time of specific recognition,opens mesoporous pore channel,activates DOX release,and serve as a"switch"for intelligent response to drug release.MSNs/DOX-APT,a tumor cell molecular tag specific recognition-trigger drug delivery system,was finally prepared through the above process.In this study,the successful preparation of MSNs and MSNs-N₃ was demonstrated by transmission electron microscope?TEM?and X-ray diffractometer?XRD?.The results of transform infrared spectroscopy?FT-IR?and fluorescence spectra showed that the hairpin APT was successfully attached to MSNs-N₃ surface.Nitrogen adsorption and desorption results showed that the specific surface area of MSNs-N₃nanoparticles was 782.10 m²/g,and the average pore diameter was 2.31 nm,which were suitable for DOX loading.The drug loading rate was determined by high performance liquid chromatography?HPLC?.It was proved that the nano-system MSNs-N₃ could efficiently load DOX,and MSNs/DOX-APT was successfully prepared with a drug loading rate of 66%.In this paper,human breast cancer?MCF-7?cells were used as model cells and human normal breast cells were as control cells to evaluate the anti-tumor activity and the selectivity of MSNs/DOX-APT in vitro.As discovered from the images of confocal laser scanning microscope?CLSM?,APT could effectively identify MCF-7 cells with high expression of MUC-1 protein and light up fluorescence with conformational changes,while it could not identify Hs578bst cells.Therefore,the drug release results of the nano-system in the two kinds of cells also showed that MSNs/DOX-APT could selectively identify MCF-7 cells and release a large number of drugs,but hardly release drugs in Hs578bst cells.The cell viability of MSNs/DOX-APT?DOX concentration of 5?g/mL?interacting with MCF-7 cells and Hs578bst cells for 72 h was 24.8%and 86.0%,respectively,indicating that the nanosystem had a selective killing effect on tumor cells.The cell experiment results showed that the MSNs/DOX-APT nanosystem could selectively activate the treatment of tumor cells and reduce the toxic side effects on normal cells.To evaluate the anti-tumor activity of the delivery system in vivo,MCF-7 cells were used to establish the tumor-bearing nude mouse model.The distribution of MSNs/IR783-APT in nude mice was investigated through in vivo imaging of small animals,and the results showed that MSNs/IR783-APT group with the recognition ability of APT could be rapidly and effectively enriched and retained at the tumor site for a long time.The pharmacodynamics results showed that the tumor volume of MSNs/DOX-APT group decreased by 1.5 times compared with the DOX group,indicating that the nano-system could be activated by tumor cells and activate the chemotherapy.The pathological results further proved the chemotherapy effect of MSNs/DOX-APT,and the reduced toxic side effects of DOX on normal tissues.