Furin Enzyme-Instructed Self-Aggregation Of Fe₃O₄ Nanoparticles For Enhanced MR T2 Imaging And Photothermal Therapy Of Tumors

The treatment of cancer as a medical challenge and research hotspots has been continuing.Therefore,effective local treatment has become the direction of research in China and abroad.As an effective and noninvasive local therapy,photothermal therapy?PTT?has become one of the new hot topics in tumor research.Fe₃O₄ NPs?Nano Particles?,as a photothermal reagent with the function of MRI T2 contrast agent,were widely used in the basic research of PTT.However,tumor cells are non-specialized phagocytes,which mainly uptake Fe₃O₄ NPs through cytosol mediated by grid protein,with a low uptake rate.Moreover,NPs were difficult to retain in tumor due to poor delivery in the body,and resulting in low efficiency of MR T2 imaging and photothermal treatment.Therefore,in order to improve the tumor delivery efficiency of NPs to enhance the effect of tumor diagnosis and treatment,it has become an urgent problem to be solved in the current biological nanomaterials research.In this study,the Ac-Arg-Val-Arg-Arg-Cys?St Bu?-Lys-CBT?RVRRCK-CBT?probe?Compound 1?was designed and combined with superparamagnetic iron oxide?SPIO?to form SPIO@1NPs.SPIO@1NPs mainly use furin enzyme and GSH to overexpress in most cancer cells.The disulfide bond of the Cys motif is reduced by high GSH,and the RVRR substrate is cleaved by furin.Instantly,the cyano groups of the CBT motifs and the free 1,2-aminothiol groups undergo a condensation reaction to form a more hydrophobic dimer,resulting in SPIO@1NPs aggregate by cross-linking in the tumor to solve the problem of low NPs tumor delivery efficiency.And the aggregation state of SPIO@1NPs can effectively improve the MR r2 relaxation efficiency and photothermal conversion efficiency to achieve the purpose of further enhancing tumor MR T2 imaging and photothermal treatment.In the first chapter,RVRRCK-CBT was synthesized and purified by high performance liquid chromatograph?HPLC?and characterized.The surface carboxylated SPIO was then coupled with the compound 1 to form SPIO@1NPs.After GSH reduction and furin cleaved,compound 1 effectively formed a dimer and was verified by HPLC and high resolution mass spectrometry?HR-MS?.After incubation with GSH and furin,that significant aggregation of SPIO@1NPs was observed under transmission electron microscope?TEM?,and the NPs size reached 139.72± 24.69 nm by DLS?Dynamic Light Scattering?.In the second chapter,following self-aggregation in vitro by furin instructed,SPIO@1NPs not only showed redshift of absorption spectrum and photothermal thermal conversion efficiency increased,but also significantly enhanced the MR transverse relaxation rate.Furthermore,the intracellular self-aggregation of SPIO@1NPs was further verified by TEM and Prussian blue staining.The results showed that SPIO@1NPs apparently self-aggregated around or near the Golgi body in MDA-MB-468 cells,and could effectively improve the cell uptake rate.The third chapter mainly investigate the self-aggregation of SPIO@ 1NPs in vivo tumor to enhance the MR T2 imaging.The self-aggregation of SPIO@1NPs in MDA-MB-468 tumor significantly improves the delivery efficiency,and enhanced MR T2 imaging,which has higher ?SNR and lower T/N ratio compared with naked SPIO NPs.Moreover,the peak time of NPs tumor accumulation provided by MR T2 imaging.The fourth chapter mainly investigate the self-aggregation of SPIO@1NPs to enhance PTT both in vitro and in vivo.It was found that SPIO@1NPs significantly increased thermal damage efficiency of cells in vitro.In vivo,pathologic evaluation of mice tumors further confirmed that SPIO@1NPs can effectively enhance PTT after self-aggregation in tumors.[Conclusion] In short,SPIO@1NPs was successfully synthesized in this study,which can significantly improve the delivery efficiency of NPs in tumor through selfaggregation instructed by furin enzyme.Moreover,the NPs in the aggregated state was used to enhance the efficiency of MR T2 imaging and photothermal therapy,effectively killing cancer cells.We believe that this basic technique,based on tumor-specific enzymeinstructed intracellular self-aggregation of NPs,could be useful for the rational synthesis of other inorganic NPs for use in the fields of tumor diagnosis and treatment.