Visualization of protrusion-localized STAT3 mRNA using a self-powered lipidic nanoflare for predicting hepatocellular carcinoma metastasis

Cancer cell metastasis is one of the major causes of patients death with hepatocellular carcinoma (HCC). Previous findings demonstrated that protrusion-accumulated STAT3 mRNA is highly related to HCC cell metastasis, making protrusion-localized STAT3 mRNA an ideal biomarker for evaluating HCC cell initiation and progression. A self-powered lipidic nanoflare (SLNF) has been developed for detecting the expression level of protrusion-accumulated STAT3 mRNA in individual HCC cells, which enables accurate prediction of HCC metastasis. The LNF system is a cholesterol micelle decorated with two kinds of DNA probes, a double-stranded response DNA and a single-stranded fuel probe. The cholesterol micelle can be easily assembled from an amphipathic cholesterol-conjugated DNA via hydrophobicity-mediated aggregation, exhibiting a highly efficient cell internalization. Moreover, the compact and high-density arrangement of DNA probes on the surface of cholesterol micelle enhances their biostability. All the above features make the LNF system an ideal approach for intracellular RNA imaging. The assay commences with the binding of STAT3 mRNA to the response DNA, which peels off the waste DNA and exposes the toehold domain. This domain serves as the proximal holding point for the fuel probe to initiate a strand displacement amplification, which is a crucial step in enabling the detection of targets expressed at trace levels, yielding a limit of detection (LOD) of 100 pM at 37 °C within 1.5 h. The SLNF system is expected to provide useful insight into the development of simple and degradation-resistant DNA probes for visual prediction of HCC metastasis, showing potential applications in tumor diagnosis and treatment.