A Multicolor Nanoprobe for Detection and Imaging of Tumor‐Related mRNAs in Living Cells

Abstract

Cancer is a leading cause of death worldwide and accounts for several millions of deaths every year. The survival of cancer patients is strongly associated with the stage of the tumor at the time of diagnosis. Identifying the cancer at the cellular level in an early stage before metastasis holds great promise for increasing the survival of cancer patients. A major focus of research towards this goal is on the estimation of abnormalities in gene expression in living cells. Tumor-related mRNA has been widely used as a specific marker to assess the migration of tumor cells locally or in the bloodstream. Changes in the level of tumor-related mRNA expression are correlated with tumor burden and malignant progression. The detection of tumor-related mRNA markers in intact cancer cells provides new tools for identifying cancer cells in clinical samples. Recently, a variety of techniques have been exploited to monitor tumor-related mRNAs. Among these methods, fluorescence imaging analysis offers an appealing approach for the detection of cancer at the cellular level, which may be of prognostic significance. Many fluorescence probes have been synthesized for the detection and imaging of mRNA in cancer cells and most of the research focuses on detecting a single type of mRNA, which may yield false positive results and limits the development of intracellular mRNA imaging and detection. Notably, cancer is associated with multiple tumor-related mRNAs, and some mRNA markers are expressed in normal cells. Simultaneous detection of multiple targets brings new opportunities for improving the accuracy of early cancer detection over the single-marker assay. Although various types of platforms for the detection of multiple targets have been developed, none have been designed for imaging three or more markers in living cells. Such intracellular imaging techniques for multiple tumorrelated mRNAs could promote the progress of early cancer detection. Herein, we describe a multicolor fluorescence nanoprobe based on nanoflares, which simultaneously detects three intracellular tumor-related mRNAs. The nanoprobe consists of gold nanoparticles (Au NPs) functionalized with a dense shell of recognition sequences (synthetic oligonucleotides) hybridized to three short dye-terminated reporter sequences by gold–thiol bond formation (Figure 1). The recognition sequences contain 21-base recognition elements for three specific mRNA transcripts: c-myc mRNA, TK1 mRNA, and GalNAc-T mRNA.