Multifunctional Bio-Nanoprobes For Cancer Diagnosis And Drug Delivery

Nanomaterials have attracted great attention in the field of biology, medicine, energy, catalysis, and environment, due to their small sizes, large surface areas, modifiable surface, unique physical and chemical properties, excellent biocompatibility, etc. Particularly, many efforts have been performed to construct multifunctional nanoprobes and nanocarriers by the combination of nanoparticles and biomolecules in fluorescent labeling, magnetic separation, gene transfection, drug delivery, and cancer therapy. In this dissertation, we have explored to choose protein receptors and microRNA overexpressed in tumor cells as targets, and fabricate multifunctional bio-nanoprobes with nanomaterials and biomolecules to realize specific fluorescent labeling, targeted drug delivery, and controlled release. The main contents are as follows:1. Multi-Shell Structured Fluorescent-Magnetic Nanoprobe for Target Cell Imaging and On-Chip SortingPolydopamine is one of the most popular species in molecular modification not only for its convenience in combination with inorganic particles but also for its high reactivity with nucleophiles such as amine and thiol in alkaline medium. Herein, we have developed a core-triple-shell structured multi-functional nanoprobe Fe3O4/SiO2/CdSeTe@ZnS-SiO2/ polydopamine with strong fluorescence and a fast magnetic response for specifically recognizing, fluorescently labeling, and magnetically sorting target tumor cells on a microfluidic chip. The outer polydopamine layer not only effectively alleviated the quenching effect of the interlayer quantum dots but also provided a convenient and versatile functional interface to readily conjugate with the recognizing model molecules of aptamer KH1C12 with amine, thiol, or carboxyl groups. Moreover, the polydopamine isolation and PEG decoration equipped the as-fabricated nanoprobes with little cytotoxicity and nonspecific affinity, leading to the effective and specific profiling of the protein epitopes expressed on the target tumor cells. Taking advantage of the magnetic property and specific recognition, the modified nanoprobe was utilized to label and isolate HL-60 cells from a homogeneous cell mixture of HL-60 and K562 cells on a microfluidic chip. This study presents an innovative strategy for developing highly integrated nanoprobes of strong fluorescence and magnetic controllability, opening up a promising probe-based avenue for biological imaging and separation.2. DNA-Hybrid-Gated Multifunctional Mesoporous Silica Nanocarriers for Dual-Targeted and MicroRNA-Responsive Controlled Drug DeliveryThe design of an ideal drug delivery system with targeted recognition and zero premature release, especially controlled and specific release that is triggered by an exclusive endogenous stimulus, is a great challenge. A traceable and aptamer-targeted drug nanocarrier has now been developed; the nanocarrier was obtained by capping mesoporous silica-coated quantum dots with a programmable DNA hybrid, and the drug release was controlled by microRNA. Once the nanocarriers had been delivered into HeLa cells by aptamer-mediated recognition and endocytosis, the overexpressed endogenous miR-21 served as an exclusive key to unlock the nanocarriers by competitive hybridization with the DNA hybrid, which led to a sustained lethality of the HeLa cells. If microRNA that is exclusively expressed in specific pathological cell was screened, a combination of chemotherapy and gene therapy should pave the way for a targeted and personalized treatment of human diseases.3. In situ Amplification of Intracellular MicroRNA with MNAzyme Nanodevices for Multiplexed Imaging, Logic Operation, and Controlled Drug ReleaseMicroRNAs (miRNAs) have participated in many biological processes, including cancer initiation, progression, and metastasis, indicative of potential diagnostic biomarkers and therapeutic targets. To tackle the low abundance of miRNAs in cells, MNAzymes (multicomponent nucleic acid enzymes) were used in constructing multifunctional nanodevices for stringent recognition and in situ signal amplification with gold nanorods as photothermal heater. The fluorescent turn-on nanodevices presented excellent specificity and sensitivity in visualizing trace amounts of miRNAs in single cell, allowing logic operation for graded cancer risk assessment and dynamic monitoring of therapy response. Meanwhile, through general molecular design, the aptamer-modified nanocarricrs loaded with doxorubicin exclusively entered the target tumor cells, where drug release rates were spatial-temporally controlled by the modulation of endogenous miRNA expression. Integrated with miRNA profiling techniques, the designed nanodevices can provide general strategy for disease diagnosis, prognosis, and combination treatment with chemotherapy and gene therapy.4. Smart Thermal-Responsive Nanocarriers for NIR-Guided Delivery and microRNA/ATP-Controlled Release of siRNA and Dox in vivoIn order to achieve the targeted delivery and controlled release of siRNA and Dox in tumor cells in vivo, smart nanocarriers, whose surface composition could be reversibly adjusted under NIR irradiation, were constructed by assembling the Y-motifs, two temperature-sensitive polymers onto the surface of gold nanorods (GRs); the PLK1 specific siRNA (siPLK1) was coupled at the end of the DNA strands, while Dox was intercalated into he GC base pairs at the Y-motif scaffold. After intravenous injection, the PEG corona around the nanocarriers efficiently protected the siRNA coupled Y-motifs from nuclease degradation and renal clearance, and favored their accumulation at tumor sites as a result of EPR effect. Under the NIR irradiation, the surface composition was altered by heating the nanocarriers through photothermal conversion, concomitant with the shrinkage of P39PEG and the exposure of the RGD shell, which specifically recognized the αvβ3 integrins overexpressed in most cancer cells, and thereby enhanced the uptake via receptor-mediated endocytosis. With the aid of heating, the nanocarriers rapidly escaped from the endosome into cytosol, where the endogenous miRNA triggered the dissociation of the Y-motifs in a high level of ATP, thereby yielding the release of intercalating Dox and coupling siRNA, which eventually activate the apoptosis-induced cell death. The smart nanocarriers have explored a new avenue for synergetic treatment of cancer with gene and drugs in a controlled manner.