Click Chemistry-Driven Dual Nanosystem for miR-122 Detection with Single-Base Specificity from Tumour Cells

MicroRNAs (miRNAs) are considered potential biomarkers due to their specific expression patterns in tissues and their changes in expression under pathological conditions. MicroRNA-122 (miR-122) is a vertebrate-specific miRNA known for its high expression in the liver, where it plays a role in hepatic metabolism, liver development and cellular differentiation. Dysregulation of miR-122 has been found in pathological conditions such as hepatocellular carcinoma or drug-induced liver injury (DILI). Currently, miRNA detection faces challenges such as improving point-of-care efficiency, obtaining direct results and accurately identifying miRNA isoforms. Overcoming these challenges is essential to improve the clinical efficacy of miR-122 detection as a diagnostic and prognostic biomarker in liver-related diseases. In this study, we present a click chemistry driven dual nanosystem designed for single-base specific miR-122 detection using a flow cytometer as the reading platform. Using the dual nanosystem, we analysed miR-122 expression in various tumour cell lines with specificity and robustness, showing a high correlation with results obtained from a TaqMan assay. We also detected miR-122 using a serum matrix and identified four single nucleotide variations at a specific position within the miR-122 sequence. Our approach involves a fully bioorthogonal strategy and includes signal amplification using two types of different nanoparticles together with the high affinity of peptide nucleic acid probes. The chemistry used in this dual nanosystem offers remarkable versatility and provides a promising opportunity to pioneer enzyme-free nanoparticle-based strategies based on click chemistry and bioorthogonality miRNA detection and isomiR studies, thus advancing the field of molecular diagnostics.