Construction Of Novel Biofunctional Nanomaterials And Their Applications In Tumor Marker Detection And Targeted Therap

Novel biofunctional nanomaterials with unique physicochemical properties,have been widely applied to biosensors,nanodrug carriers,and microdevices,exhibiting unique advantages in biochemical analysis,early disease diagnosis and therapy.Metal organic frameworks（MOFs）have been developed into a new class of functional materials with great potential as nanocarriers to effectively improve the loading and utilization of biomolecules due to their excellent specific surface areas,unique pore structures.In addition,DNA as a biomaterial has been widely used in biosensing,bioimaging and biomedicine owing to its good biocompatibility,sequence programmability,adjustable size and structure.In this paper,combining zeolitic imidazole framework（ZIF-8）material with a three-dimensional DNA walker,a novel electrochemical biosensor was constructed,achieving label-free exponential signal amplificated detection of tumor marker micro RNA-21（mi RNA-21）.Further,functional flower-like DNA nanomaterials which loaded with glucose oxidase（GOx）and chemotherapeutic drug doxorubicin（DOX）were synthesized based on copper ion(Cu²⁺)mediated rolling circle amplification（RCA）technique and had been successfully applied to chemo/starvation therapy（ST）/chemodynamic therapy（CDT）trimodal combination therapy for cervical cancer.This work provides new methods and ideas for the research and application of new bionanomaterials in the fields of cancer diagnosis and treatment.The thesis contains the following two works:1.Metal-organic framework nanoreactor-based electrochemical biosensor coupled with three-dimensional DNA walker for label-free detection of micro RNAIn this work,an electrochemical biosensor integrating the advantages of ZIF-8nanoreactor and 3D DNA walker was constructed for the detection of mi RNA-21.GOx@ZIF-8 was synthesized by a one pot method,and then loaded with gold nanoparticles（Au NPs）by the aid of electrostatic adsorption to form Au NPs/GOx@ZIF-8nanoreactor which was used to modify the electrode.The Au NPs as the support was employed to constructed DNA orbit by modifying the H1 on its surface.3D DNA walker was triggered by target mi RNA-21 to generate DNA mimicking enzyme（HRP DNAzyme）with horseradish peroxidase activity at the surface of Au NPs/GOx@ZIF-8via strand substitution reaction with fuel DNA and track DNA（H2）.Meanwhile,the cycling of mi RNA-21 was realized,which enhanced the cascade catalytic reaction between GOx and HRP DNAzyme and improved the electrochemical signal.As an orbital support for 3D DNA walkers,Au NPs also played the role of a bridge for electron transport,further improving the sensitivity.This proposed electrochemical biosensor with high specificity exhibited an excellent performance for amplification detection of mi RNA-21 in the dynamic working range from 0.1 n M to 10μM with a detection limit of 29 p M,and was successfully applied to real sample detection of mi RNA-21,showing potential prospects in clinical diagnostic and opening up a broad avenue for the development of DNA nanotechnology in the field of biosensing.2.Novel DNA nanoflowers based on co-loading of glucose oxidase and DOX for the treatment of cervical cancerIn this work,a novel DNA flower nanomaterial DOX/GOx@CuDNFs was synthesized by combining the liquid crystallization of long-chain products of RCA reaction and the coordination between copper ions(Cu²⁺)and DNA molecules,achieving the co-loading of GOx and chemotherapeutic drug doxorubicin（DOX）.RCA template was designed to contain the complementary sequence of Hela cell nucleic acid aptamer as well as that of DOX DNA loading sequence.The Cu²⁺-mediated RCA reaction was performed by one-pot method to generate CuDNFs with the GOx encapsulation at the same time,and the formed GOx@CuDNFs was further incubated with DOX to produce DOX/GOx@CuDNFs composites.The morphology,efficiency of hydroxyl radical（·OH）generation,loading and release of DOX and catalytic activity of the prepared novel functional nanomaterials were systematically investigated.The DOX/GOx@CuDNFs exhibited excellent catalytic activity and the self-assembly ability of which could be effectively enhanced with the assistance of Cu²⁺.When DOX/GOx@CuDNFs specific recognize and entry into Hela cells,the Cu²⁺was reduced to Cu⁺by intracellular high concentration glutathione（GSH）,causing the disassembly of DOX/GOx@CuDNFs to release DOX.GOx could catalyze intracellular glucose to produce H₂O₂which was catalyzed by Cu⁺to generate·OH,thus realizing the chemo/starvation/chemodynamic trimodal synergistic targeted therapy for cervical cancer,providing new approaches and thinkings for DNA nanotechnology as well as combined diagnosis and treatment of cancer.