Construction Of Multifunctional Nanoparticles Based On Ferritin And Its Application In The Integration Of Tumor Diagnosis And Treatment

At present,cancer is still one of the main causes of death in the world,with about one sixth of the world’s people dying of malignant tumors.Photodynamic therapy(PDT)has become an alternative or additional treatment method of chemotherapy and surgery in tumor treatment due to its high specificity,minimally invasive and controllability.The essence of PDT is to use photosensitizer(PS)to react with oxygen under light to generate reactive oxygen species(ROS),then causing apoptosis of tumor cells.In addition,PS shows inherent fluorescence after illumination,which is conducive to imaging-guided therapies.However,traditional PSs are affected by the aggregation induced quenching(ACQ).The discovery of aggregation induced emission(AIE)provides a new possibility for fluorescence imaging and PDT.AIE has the opposite properties to traditional PSs: it hardly emits light at low concentration,but it can emit strong fluorescence at high concentration or aggregation state.However,most AIE materials(AIEgens)need to be wrapped into nanocomposites to improve their biocompatibility and tumor targeting.In this study,the gene of human ferritin heavy chain(HFtn)was first cloned into p GEX-6p-1 vector,and then the tumor homing and penetrating peptide LinTT1 was inserted into the N-terminal of HFtn by gene mutation,and then the vector was transformed into E.coli.After induction of expression,affinity chromatography,ion exchange,and gel filtration,LinTT1-HFtn with purity of more than 90% was obtained.Me TTMN,an AIE material,was encapsulated in LinTT1-HFtn nanocages via the mean of p H-dependent disassembly/reassembly,thereby synthesizing a nano-system(LinTT1-HFtn-AIE NPs)that can target tumor,enable fluorescent imaging and achieve photodynamic therapy.Then,a human hepatoma cell line Hep G2 was used to construct an in vitro tumor cell sphere model to determine the localization of LinTT1-HFtn-AIE NPs in Hep G2 cells,the ability to produce ROS,the penetration ability in the tumor cell sphere model in vitro,and the damage to mitochondria and Hep G2 cells.In addition,LinTT1-HFtn-AIE NPs were injected into tumor(Hep G2)-bearing mice through tail vein injection to test the tumor-targeting ability and therapeutic effect of LinTT1-HFtn-AIE NPs in mice in vivo.The results showed that LinTT1-HFtn-AIE NPs had good biological safety,excellent water solubility,good stability and uniform particle size distribution.Due to the respective penetration and targeting effects of LinTT1 and HFtn,LinTT1-HFtn-AIE NPs could successfully target into the mitochondria of tumor cells,and realize emergent image guidance for tumor treatment.In addition,no matter the LinTT1-HFtn-AIE NPs were free outside of cells,in the tumor cells,or in the mice body,the NPs had the ability to produce a large amount of ROS under white light.ROS can cause mitochondrial damage,tumor cell death,and ultimately achieve the goal of photodynamic therapy of tumor.Meanwhile,PDT,as a novel cancer therapy,is gaining popularity due to its high specificity,controllability and minimal invasiveness.Our preliminary experiment also shows that LinTT1-HFtn-AIE NPs are of great significance for the development of clinical photodynamic therapy.In addition,to further investigate the potential application of ferritin nanoplatform in the treatment of triple-negative breast cancer(TNBC),bioinformatics research on TNBC was also carried out in this paper,with a view to finding potential therapeutic targets,which provide the research basis for constructing ferritin nanoparticles specifically targeting TNBC.More and more evidences show that glycolysis plays an important role in tumor pathogenesis,invasion,chemotherapy resistance,metastasis and immune escape.However,there is still a lack of all-round analysis on the diagnosis and prognostic significance of glycolysis in TNBC.The second part of this dissertation work constructed the risk characteristics related to glycolysis by analyzing the transcriptomics and clinical data of TNBC patients.First,the transcriptome and clinical data of TNBC patients were collected from the database of the Molecular Taxonomy of Breast Cancer International Consortium(METABRIC)and The Cancer Genome Atlas(TCGA);Glycolysis-related genes(GRGs)were obtained from the Molecular Signatures Database(MSig DB).Then,the differentially expressed GRGs(DE-GRGs)related to TNBC were obtained by differential comparative analysis.Finally,based on the DE-GRGs,the risk characteristics related to glycolysis were established by using Least Absolute Shrinkage and Selector Operation(LASSO)and multivariate Cox regression analysis.The tumor microenvironment,mutation status,prognostic value and chemotherapy response of different risk groups were analyzed,and the external validation was analyzed using the independent cohort of METABRIC database.The results showed that the prognostic characteristics related to glycolysis included five genes(IFNG,IRS2,ACSS2,GAL3ST1 and GFUS),which could predict the prognosis of patients with TNBC independently of clinical factors(p<0.05).The patients were divided into low-risk group and high-risk group according to the median risk score.Compared with the low-risk group of TNBC patients,the total survival period of high-risk patients was significantly reduced(HR=2.718,p<0.001).Receiver operating characteristic and calibration curves also showed that the model had high performance in predicting risk stratification and survival rate.After external verification,the results were consistent with the experimental results.Therefore,the glycolysis and prognosis-related(GRP)feature based on GRGs can predict the prognosis of patients with TNBC and may be helpful for clinical decision-making of these patients.