Application Of Dual-responsive Upconversion Nanoplatform In Tumor-targeted Imaging And Therapy

Objective:The diagnosis and treatment of cancer at an early stage are of great significance for prolonging the survival time and improving the life quality of patients.Theranostic nanoplatforms integrating the functions of bioimaging,targeted delivery and controlled release of therapy agents into a single nanoparticle enable simultaneous diagnosis and therapy of tumor.Imaging-guided tumor therapy can reduce side effects,increase therapeutic efficiency.The unique up-conversion luminescence(UCL)properties of rare earth ion-doped up-conversion nanoparticles(UCNPs)and the application for photodynamic therapy,photothermal therapy and drug delivery make them promising candidates for theranostic nanoplatform construction.Herein,a cis-diol/pH dualresponsive UCNPs-based theranostic platform loaded with antitumor drug camptothecin(CPT)has been firstly developed for cancer imaging and therapy.The dual-responsive property of borate ester bond can further enhance the drug release at tumor site,improve the therapeutic efficiency and provide a new approach for the functional modification of UCNPs-based drug delivery systems and the controlled release of drugs in tumor cells.Content:The research synthesized hyaluronic acid-modified up-conversion nanoparticles.CPT,an anti-tumor drug was bonded to the surface of nanoparticles using dualresponsive borate ester bond to produce tumor-targeted theranostic nanoplatform.We studied its drug release behavior,cell targeting ability,intracellular localization and cytotoxicity,and preliminarily investigated its distribution in vivo,metabolism behavior,and imaging ability.Methods:1.The synthesis and characterization of CPT-UCNPs.First,the hyaluronic acid(HA)modified UCNPs(HA-UCNPs)were synthesized by solvothermal method.Based on the specific affinity of HA and CD44 receptors,HA-UCNPs could target tumor cells with high expression of CD44 receptors.Aminobenzeneboronic acid(APBA)was modified on the surface of HA-UCNPs through amide condensation reaction to obtain APBA-UCNPs,having the capacity for bonding therapeutics containing cis-diol structure.The cis-diol structure was introduced to 10-hydroxycamptothecin using 3-(3,4-dihydroxyphenyl)propionic acid(DHPA)to obtain CPT-DHPA.CPT-UCNPs were fabricated based on the covalent binding of CPT-DHPA onto APBA-UCNPs.The physicochemical properties of CPT-UCNPs were characterized by transmission electron microscope(TEM),particle size analyzer,fluorescence spectrophotometer and infrared spectrometer,etc.2.In vitro drug release behavior of CPT-UCNPs.Drug release behavior in vitro was evaluated by dialysis method by simulating the environment of normal tissues,tumor tissues and lysosomes.3.The cell targeting ability,intracellular localization and cytotoxicity of CPTUCNPs.Tumor cells with different expression levels of CD44 receptors were used to study the cell targeting ability,intracellular localization and cytotoxicity of CPTUCNPs through cell uptake assay and MTT cell proliferation inhibition assay.4.In vivo metabolism behavior,distribution and imaging of CPT-UCNPs.The in vivo metabolism behavior,distribution and imaging ability of CPT-UCNPs in mice were studied by intravenous injection to investigate the potential application of the nanoprobe.Results:1.The synthesis and characterization of CPT-UCNPs.The characterization results of CPT-UCNPs via TEM,fluorescence spectrophotometry and infrared spectroscopy showed that CPT-UCNPs was successfully synthesized.The prepared CPT-UCNPs was spherical,presented narrow size distribution and good colloidal stability.Under 980 nm NIR light excitation,CPT-UCNPs emitted strong multicolor UCL at 540 nm and 660 nm with good photostability.The drug loading efficiency 1.5 wt% with the optimal feeding ratio of 3:1(CPT-DHPA : APBA-UCNPs).2.In vitro drug release behavior of CPT-UCNPs.The drug release experiments in vitro demonstrated a controlled drug release behavior of CPT-UCNPs upon dualresponsiveness.CPT-UCNPs liberated drugs triggered by competitive glucose at low pH,showing a synergistic effect of pH and glucose on drug release.3.The cell targeting ability,intracellular localization and cytotoxicity of CPTUCNPs.Cell uptake assays demonstrated that CPT-UCNPs could achieve targeted drug delivery and UCL imaging of HCT116 cells through the specific recognition of HA ligands towards CD44 receptors on cells.CPT-UCNPs entered tumor cells via receptormediated endocytosis and CPT dissociated from the UCNPs in lysosomes under the acidic and glucose environment to produce dosage-dependent cytotoxic effects.4.In vivo metabolism behavior,distribution and imaging of CPT-UCNPs.CPTUCNPs were mainly distributed in the liver and spleen areas via intravenous injection,and could emit stable UCL signal excited by 980 nm NIR light with low interference from the autofluorescence induced by biological tissue,which demonstrated the potential bioimaging application in vivo.Conclusion:The CPT-UCNPs theranostic nanoprobe we designed could achieve the aim for targeted UCL imaging and drug delivery to tumor cells with high expression of CD44 receptors.CPT-UCNPs entered the endosome/lysosome pathway after being taken up by tumor cells via receptor-mediated endocytosis.The acidic environment and the presence of glucose in lysosomes triggered the dual-responsive mechanism of the borate ester bond to achieve controlled release of CPT.CPT-UCNPs with high signalto-noise ratio and benign photostability in vivo shows its potential application in bioimaging in vivo.