Amylase-Protected Ag Nanodots for in vivo Fluorescence Imaging and Photodynamic Therapy of Tumors
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Background: Fluorescent metallic nanodots (NDs) have become a promising nanoprobe for a wide range of biomedical applications. Because Ag NDs have a high tendency to be oxidized, their synthesis and storage are a big challenge. Thus, the method for preparing stable Ag NDs is urgently needed. Surface modification and functionalization can enrich the capability of Ag NDs.
Methods: In this work, fluorescent Ag NDs were synthesized in deoxygenated water by using porcine pancreatic α-amylase (PPA) as the stabilizing/capping agent. The absorption and fluorescence of PPA-protected Ag NDs (PPA@AgNDs) were measured with a spectrophotometer and a spectrofluorometer, respectively. The morphology of PPA@AgNDs was characterized by high-angle annular dark-field (HAADF) scanning transmission electron microscopy (STEM). The biocompatibility of PPA@AgNDs was evaluated by tetrazolium (MTT)-based assay. PolyLys-Cys-SH (sequence: KKKKKKC) peptides were conjugated to PPA@AgNDs via heterobifunctional crosslinkers. PolyLys-Cys-linked PPA@AgNDs absorbed 5-aminolevulinic acid (ALA) by electrostatic interaction at physiological pH. The capability of tumor targeting was evaluated by intravenously injecting PPA@AgND-ALA into 4T1 breast cancer xenograft mouse models. Photodynamic therapy (PDT) against tumors was performed under 635 nm laser irradiation.
Results: PPA@AgNDs emitted at 640 nm with quantum yield of 2.1%. The Ag NDs exhibited strong photostability over a long period and a fluorescence lifetime of 5.1 ns. PPA@AgNDs easily entered the cells to stain the nuclei, showing the capabilities of living cell imaging with negligible cytotoxicity. ALA-loaded PPA@AgNDs (PPA@AgND-ALA) presented the superiority of passive tumor targeting via the enhanced permeability and retention (EPR) effect. Tumors were visualized in the near-infrared (NIR) region with reduced background noise. ALA molecules released from PPA@AgND-ALA was converted into the photosensitizer (PS) of protoporphyrin IX (PpIX) intracellularly and intratumorally, which greatly improved the PDT efficacy.
Conclusion: Our approach opens a new way to design a novel theranostic nanoplatform of PPA@AgND-ALA for effective tumor targeting and fluorescence image-guided PDT.
Keywords: Ag nanodots; peptides; photodynamic therapy; targeted imaging; α-amylase.
