All-in-One Fusogenic\nNanoreactor for the Rapid Detection\nof Exosomal MicroRNAs for Breast Cancer Diagnosis

Molecular-profiling-based cancer diagnosis has significant\nimplications\nfor predicting disease prognosis and selecting targeted therapeutic\ninterventions. The analysis of cancer-derived extracellular vesicles\n(EVs) provides a noninvasive and sequential method to assess the molecular\nlandscape of cancer. Here, we developed an all-in-one fusogenic nanoreactor\n(FNR) encapsulating DNA-fueled molecular machines (DMMs) for the rapid\nand direct detection of EV-associated microRNAs (EV miRNAs) in a single\nstep. This platform was strategically designed to interact selectively\nwith EVs and induce membrane fusion under a specific trigger. After\nfusion, the DMMs recognized the target miRNA and initiated nonenzymatic\nsignal amplification within a well-defined reaction volume, thus producing\nan amplified fluorescent signal within 30 min. We used the FNRs to\nanalyze the unique expression levels of three EV miRNAs in various\nbiofluids, including cell culture, urine, and plasma, and obtained\nan accuracy of 86.7% in the classification of three major breast cancer\n(BC) cell lines and a diagnostic accuracy of 86.4% in the distinction\nbetween patients with cancer and healthy donors. Notably, a linear\ndiscriminant analysis revealed that increasing the number of miRNAs\nfrom one to three improved the accuracy of BC patient discrimination\nfrom 78.8 to 95.4%. Therefore, this all-in-one diagnostic platform\nperforms nondestructive EV processing and signal amplification in\none step, providing a straightforward, accurate, and effective individual\nEV miRNA analysis strategy for personalized BC treatment.