Yu‐Fen Huang

We have developed a highly specific sensing system for platelet-derived growth factors (PDGFs) and platelet-derived growth factor receptors (PDGFR) that uses gold nanoparticles (GNPs). We synthesized GNPs modified with an aptamer (Apt-GNPs) that is specific to PDGFs and used them to detect PDGFs by monitoring the changes in the color and extinction of the Apt-GNPs that occur as a result of aggregation. The color of the Apt-GNPs changes from red to purple at low concentrations (<400 nM), but changes only slightly at higher concentrations (>400 nM). We found that the sensitivity of the Apt-GNPs for the three PDGFs is highly salt-dependent, with an optimum condition of 200 mM NaCl. We obtained biphasic curves when plotting of the ratios of the extinction coefficients of the Apt-GNPs at 650 and 530 nm against the concentrations of PDGF-AA at various concentrations of Apt-GNPs. The linear ranges of the increases and decreases in this extinction ratio are 2.5-10 and 10-20 nM, respectively, for 0.42 nM Apt-GNPs and 25-75 and 75-200 nM, respectively, for 8.4 nM Apt-GNPs. When using 8.4 nM Apt-GNPs, the corresponding linear ranges of the increases and decreases in this extinction ratio are 15-100 and 100-400 nM, respectively, for PDGF-AB and 35-150 and 150-400 nM, respectively, for PDGF-BB. In addition, we have developed a homogeneous assay to detect the PDGF receptor-beta (PDGFR-beta) at concentrations as low as 3.2 nM, on the basis of the competition between the Apt-GNPs and PDGFR-beta for PDGF-BB. The results we present in this paper imply that there are practical applications of Apt-GNPs in protein analysis and cancer diagnosis.

Abstract Special delivery! An aptamer‐directed anticancer drug was molecularly engineered to be delivered to target cells for efficient therapeutic application. The covalent conjugation of drug and aptamer creates alternative opportunities for targeted therapy, as multiple yet specific aptamers can be “generated” relatively easily by cell‐SELEX for any target cells; this demonstrates the full potential of cell‐SELEX as a molecular discovery tool for biomedical studies and drug development. magnified image The conjugation of antitumor drugs to targeting reagents such as antibodies is a promising method that can increase the efficacy of chemotherapy and reduce the overall toxicity of the drugs. In this study, we covalently link the antitumor agent doxorubicin (Dox) to the DNA aptamer sgc8c, which was selected by the cell‐SELEX method. In doing so, we expected that this sgc8c–Dox conjugate would specifically kill the target CCRF‐CEM (T‐cell acute lymphoblastic leukemia, T‐cell ALL) cells, but with minimal toxicity towards nontarget cells. The results demonstrated that the sgc8c–Dox conjugate possesses many of the properties of the sgc8c aptamer, including high binding affinity (K d =2.0±0.2 n M ) and the capability to be efficiently internalized by target cells. Moreover, due to the specific conjugation method, the acid‐labile linkage connecting the sgc8c–Dox conjugate can be cleaved inside the acidic endosomal environment. Cell viability tests demonstrate that the sgc8c–Dox conjugates not only possess potency similar to unconjugated Dox, but also have the required molecular specificity that is lacking in most current targeted drug delivery strategies. Furthermore, we found that nonspecific uptake of membrane‐permeable Dox to nontarget cell lines could also be inhibited by linking the drug with the aptamer; thus, the conjugates are selective for cells that express higher amounts of target proteins. Compared to the less effective Dox‐immunoconjugates, these sgc8c–Dox conjugates make targeted chemotherapy more feasible with drugs having various potencies. When combined with the large number of recently created DNA aptamers that specifically target a wide variety of cancer cells, this drug‐aptoconjugation method will have broad implications for targeted drug delivery.

Molecular recognition toward specific cells is a key issue for effective disease, such as cancer, diagnosis and therapy. Although many molecular probes such as aptamers and antibodies can recognize the unique molecular signatures of cancer cells, some of these probes only have relatively weak binding affinities. This results in poor signaling and hinders cell targeting. Here, we use Au-Ag nanorods (NRs) as a nanoplatform for multivalent binding by multiple aptamers on the rod to increase both the signal and binding strengths of these aptamers in cancer cell recognition. Up to 80 fluorophore-labeled aptamers can be attached on a 12 nm x 56 nm NR, resulting in a much stronger fluorescence signal than that of an individual dye-labeled aptamer probe. The molecular assembly of aptamers on the NR surfaces also significantly improves the binding affinity with cancer cells through simultaneous multivalent interactions with the cell membrane receptors. This leads to an affinity at least 26-fold higher than the intrinsic affinity of the original aptamer probes. As determined by flow cytometric measurements, an enhancement in fluorescence signal in excess of 300-fold is obtained for the NR-aptamer-labeled cells compared with those labeled by individual aptamer probes. Therefore, the molecular assembly of aptamers clearly shows potential applications for the elucidation of cells with low density of binding sites, or with relatively weak binding probes, and can thus greatly improve our ability to perform cellular imaging and targeting. This is an excellent example of using nanomaterials to develop advanced molecular binders with greatly improved properties for cellular studies.

A near-infrared light-responsive drug delivery platform based on Au-Ag nanorods (Au-Ag NRs) coated with DNA cross-linked polymeric shells was constructed. DNA complementarity has been applied to develop a polyacrylamide-based sol-gel transition system to encapsulate anticancer drugs into the gel scaffold. The Au-Ag NR-based nanogels can also be readily functionalized with targeting moieties, such as aptamers, for specific recognition of tumor cells. When exposed to NIR irradiation, the photothermal effect of the Au-Ag NRs leads to a rapid rise in the temperature of the surrounding gel, resulting in the fast release of the encapsulated payload with high controllability. In vitro study confirmed that aptamer-functionalized nanogels can be used as drug carriers for targeted drug delivery with remote control capability by NIR light with high spatial/temporal resolution.