| Background and ObjectiveLiver cancer is one of the most common malignant tumors in the world,which seriously affects human life,health and safety.It is of great significance to realize effective treatment of liver cancer.Currently,chemotherapy is the most commonly used method to treat liver cancer,especially in advanced liver cancer.However,chemotherapy as a treatment for systemic administration is often limited by the inherent properties of traditional chemotherapeutic molecules.For example,chemotherapeutic molecules have no selectivity and specificity for killing.After they are transported throughout the body through the blood circulation,they can affect healthy tissue And cells will produce unpredictable toxic and side effects;at the same time,chemotherapeutic drugs have small molecular weights,poor biocompatibility,short residence time at the tumor site,and cannot achieve effective blood drug concentrations.Large doses or multiple administrations are needed to achieve therapeutic effects.Cause tumor multi-drug resistance and so on.Therefore,the design of intelligent drug carriers that can target tumor cells and controllable release of drug molecules,thereby achieving precise treatment of tumors,has important application value while effectively reducing toxic side effects during chemotherapy and improving the utilization efficiency of drug molecules.Aptamer is a single-stranded DNA or RNA with strong target binding ability screened by SELEX technology.It has the advantages of good biocompatibility,easy preparation and storage,good tissue permeability,and no immunogenicity.It is called As a "chemical antibody",it is a tumor-targeting molecule with great potential.I-motif is a DNA sequence rich in cytosine nucleotides,which can realize the transformation of different configurations at different p H values.It is a widely used intelligent control element.In this study,we plan to use the aptamer tumor targeting properties,combined with its soft structure and strong plasticity,to design a cleaved aptamer specifically targeting SMMC-7721 liver cancer cells,and then adapt the cleaved nucleic acid to After the ligand hybridizes with a short strand of DNA,the chemotherapeutic molecule Adriamycin(DOX)is loaded,and the I-motif sequence is further combined as an intelligent control element to construct a split aptamer-based The intelligent "nanocapsule" utilizes the slightly acidic characteristics of the tumor microenvironment to achieve specific targeting and intelligent drug release of SMMC-7721 liver cancer cells,and further explore the ability of this "nanocapsule" to grow,migrate and invade liver cancer cells The impact of this technology provides new strategies for the precise treatment of clinical tumors.Materials and methods1.Construction of smart “nanocapsules” based on split aptamers: Two fragments of split aptamers that specifically target SMMC-7721 liver cancer cells are passed through a DNA sequence with drug-carrying capabilities(ie,carrying The drug device)is connected,and the DOX can be loaded by adding the complementary sequence of the drug carrier to realize the construction of the cleaved nucleic acid aptamer "nanocapsule" with the targeting ability of SMMC-7721 liver cancer cells;then the cleaved nucleic acid One end of the aptamer sequence is hybridized with a DNA sequence(ie,a controller)containing I-motif to construct a smart "nanocapsule" that is regulated by the cell microenvironment and has the ability to target SMMC-7721 liver cancer cells.Through flow cytometry,laser confocal,fluorescence spectrophotometer,etc.,the sequence lengths of connectors and controllers are optimized and screened to obtain "nanocapsules" with high selectivity,high specificity,and high intelligence.2.Intelligent "nanocapsule" drug loading and controlled release: simply mix the chemotherapeutic drug DOX and "nanocapsule" in a solution to achieve drug loading.Use a fluorescence spectrophotometer to investigate the drug loading capacity and stability of the "nanocapsule" Sex.Then,the targeted drug delivery ability of "nanocapsules" was investigated by flow cytometry and laser confocal.3.Smart "nanocapsules" inhibit the growth,migration and invasion of SMMC-7721 liver cancer cells: use methods such as MTT,Annexin V-FITC/PI,caspase-3,cell viability determination to test the inhibitory effect of smart "nanocapsules" on the growth of SMMC-7721 liver cancer cells;use the scratch test to detect the effect on cell migration;Transwell Experiment to detect the influence on cell invasion ability.Results1.Successfully constructed nanocapsule ZI13.2.Successfully constructed nanocapsule ZB5.3.Successfully constructed the intelligent "nanocapsule" ZBI5-DOX based on split aptamer.4."Nanocapsule" ZBI5-DOX has the specific targeting and intelligent drug release capabilities of SMMC-7721 liver cancer cells.5."Nanocapsule" ZBI5-DOX can inhibit the growth,migration and invasion of SMMC-7721 liver cancer cells to a certain extent.ConclusionThe smart “nanocapsule” ZBI5-DOX based on split aptamer can be effectively activated in the tumor microenvironment,has the specific targeting and intelligent drug release capabilities of SMMC-7721 liver cancer cells,and can inhibit SMMC-7721 liver cancer cell growth,migration and invasion. |
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