Aptamer-functionalized Composite Metal Nanomaterials For Cancer Theranostics

Theranostics is a new treatment strategy combining diagnosis with therapy, which is regarded as the mainstream of treating many diseases in the future and is crucial to the achievement of personalized medicine. It is of great importance to design and synthesize high target, high sensitivity, high efficacy, high stability and low toxic side effect of theranostic multi-functional probe for the the development and application of theranostics techniques. With the emergenc e and rapid development of nanotechnology, a series of functional nanomaterials with unique physical and chemical properties have been developed, providing an opp ortunity for the construction of theranostic multi-functional probe. Meanwhile, as chemical ―antibody‖, Aptamer holds advantages of low molecular weight, low immunogenicity, easy chemical synthesis and labeling, wide range of target molecules, flexible and diverse sequence design and so on. Especially for the complex in vivo system, the tumor specific Aptamer provides an ideal kind of targeted recognition molecule to meet the demand in developing high effective theranostc probes, resulting from its properties of fast tissue penetration and uptake, short residence time in the blood and non-target organs, high target tissue accumulation rate and so on. In this thesis, aiming at the exploration of theranostic multi-functional probe, a series of novel composite metal nanomaterials have been first prepared and theranostic multi-functional probes have been subsequently designed and constructed by combination of Aptamer. The detailed description is listed as follows:1. Synthesis of Au@Ag-Au nanoparticles with broad and intense near-infrared absorption for photothermal cancer therapyAs a widely-adopted agent for photothermal therapy(PTT), Au NRs remain problematic due to the cytotoxicity derived from CTAB and the comparatively weak NIR absorption band. To address this problem, we here proposed a shape-controllable and spectrum-adjustable synthesis method of Au@Ag-Au NPs through the first coating of a Ag nanolayer outside the Au NR seed and the subsequent replacement reaction with HAu Cl4 to yield a Ag/Au alloy nanoshell. Results from TEM and UV-vis spectra analysis showed that thicknesses of Ag layers directly determined NPs’ shapes and sizes, and the formation of Ag/Au nanoshells effectively enhanced NPs’ NIR absorbance. Remarkably, the optimum Au@Ag-Au nanospheres(NSs) with a diameter of ~40 nm were revealed to have a broad and intense absorption cross section from 400 to 1100 nm, ~4.7 times higher hyperthermia effect than Au NRs, and low dark-cytotoxicity. By using A549 lung cancer as the model, a series of in vitr o investigations were performed and demonstrated that Au@Ag-Au NSs could efficaciously destruct cancer cells under a 980 nm irradiation. The PTT efficacy could be further improved by increasing NSs concentration, incubation time or irradiation time. Moreover, a preliminary in vivo study also showed that after injection into the A549 tumor, Au@Ag-Au NSs could cause an obvious necrosis at the irradiation site. It is indicated that a novel kind of promising and highly-effective NIR PTT agent has been developed, which might greatly advance the application of PTT in biomedical researches.2. Au@Ag-Au nanospheres assembled with activatable Aptamer probes for image-guided cancer thermotherapyAlthough nanomaterial-based theranostics have increased positive expectat ions for cancer treatment, it remains challenging to develop in vivo ―nano-doctors‖ affording high-contrast image-guided site-specific therapy. Here we designed an activatable theranostic nanoprobe(ATNP) via self-assembly of activatable Aptamer probes(AAPs) on Au@Ag-Au NSs. As both quenchers and heaters, Au@Ag-Au NSs were novelly prepared showing excellent fluorescence quenching and more effective near-infrared photothermal therapy than Au nanorods. The AAP comprised a thiolated Aptamer and a fluorophore-labeled complemented DNA. Thus, the ATNP with quenched fluorescence in free state could realize signal activation through target binding-induced conformational change of the AAP, and then achieve on-demand treating under image-guided irradiation. By using S6 Aptamer as the model, in vitro and in vivo studies of A549 lung cancer verified that the ATNP greatly improved imaging contrast and specific destruction, suggesting a robust and versatile theranostic strategy for the future personalized medicine.3. Aptamer-functionalized Cu-Au alloy nanostructures for in vivo cancer theranosticsAs a star material in cancer theranostics, photoresponsive gold(Au) nanostructures may still have drawbacks, such as low thermal conductivity, irradiation-induced melting effect and high cost. To solve the problem, copper(Cu) with much higher thermal conductivity and lower cost was rationally introduced to generate a novel Cu-Au alloy nanostructure(Cu-Au NS) through a simple, gentle and one-pot synthetic method. Gathering meri ts of Cu and Au, the irregularly-shaped Cu-Au NSs showed several advantages over traditional Au nanorods, including a broad and intense near-infrared(NIR) absorption band from 400 to 1100 nm, an excellent heating performance under different laser irradiat ion and even a notable photostability against melting. Then, via a facile conjugation of fluorophore-labeled Aptamers(F-Apt) on NSs, active targeting and signal output were simultaneously introduced, thus constructing a theranostic platform based on F-Apt-coated Cu-Au NSs(Cu-Au@F-Apt NSs). By using human leukemia CCRF-CEM cancer and its Aptamer Cy5-Sgc8 c as the model, a selective fluorescence imaging and NIR photothermal therapy was successfully realized for both in vitro cancer cells and in vivo tumor tissues. It was revealed that Cu-Au@Cy5-Sgc8 c NSs not only held robust target recognition and stable signal output for molecular imaging in complex biological systems, but also killed target cancer inside mice only with a 5-min 980 nm irradiation. The platform was found to be facile, stable, biocompatible and highly-effective, which might hold great potentials as a versatile tool for cancer theranostics.4. Iodide-responsive Cu-Au nanoparticle-based colorimetric platform for ultrasensitive detection of target cancer cellsColorimetric analysis is promising in developing facile, fast and point-of-care cancer diagnosis techniques, but the existing colorimetric cancer cell assays remain problematic because of dissatisfactory sensitivity as well as complex probe d esign or synthesis. To solve the problem, we here present a novel colorimetric analytical strategy based on iodide-responsive Cu-Au nanoparticles(Cu-Au NPs) combined with the iodide-catalyzed H2O2-TMB(3,3,5,5-tetramethylbenzidine) reaction system. In this strategy, bimetallic Cu-Au NPs prepared with an irregular shape and a diameter of ~15 nm could chemically absorb iodide, thus indirectly inducing colorimetric signal variation of the H2O2-TMB system. By further utilizing its property of easy biomolecule modification, a versatile colorimetric platform was constructed for detection of any target that could cause the change of Cu-Au NPs concentration via molecular recognition. As proof of concept, an analysis of human leukemia CCRF-CEM cells was performed using Aptamer Sgc8c-modified Cu-Au NPs as the colorimetric probe. Results showed that Sgc8c-modified Cu-Au NPs successfully achieved a simple, label-free, cost-effective, visualized, selective and ultrasensitive detection of cancer cells. Moreover, feasibili ty was demonstrated for cancer cell analysis in diluted serum samples. The iodide-responsive Cu-Au NP-based colorimetric strategy might not only afford a new design pattern for developing cancer cell assays, but also greatly extend the application of the i odide-catalyzed colorimetric system.5. Facile synthesis of Cu-Au-Pt alloy nanoparticles and their catalytic and plasmonic properties for cancer theranosticsThrough an integration of two or more species, the properties of complex metal nanocrystals can be significantly improved due to various synergistic effects. Herein, a facile and gentle one-pot synthetic method has been first proposed to prepare Cu-Au-Pt nanoparticles(NPs) simultaneously with excellent catalytic activity and high efficient hyperthermia. On the basis of these features, an easy-preparation, label-free theranostic probe was subsequently constructed via self-assembly of Aptamers on Cu-Au-Pt NPs and colorimetric analysis for target cells was performed using the probe. Upon irradiation by a 780 nm laser, the probe displayed a notable heating efficiency for killing target cells. Thereupon, taking advantages of multiple metal elements, Cu-Au-Pt NPs developed here might be the important supplement of theranostic materials and show a potential to provide a wide range of approaches for biosensing and other biomedical fields through utilization of different recognition elements including peptide, Aptamer and antibody.