A novel NIR absorption and emission probe for imaging of RNA dynamics in live cells and tumor models

This study presents the development of a novel RNA-targeted fluorescent probe, O-698, for real-time imaging of RNA in living cells and tumor models. O-698 exhibits significant RNA sensitivity with low cytotoxicity and strong near-infrared (NIR) fluorescence. Subcellular localization studies demonstrated that O-698 selectively binds to RNA in nucleoli and cytoplasm, with minimal overlap with other organelle markers, and outperforms existing commercial RNA probes in terms of specificity and intensity. Furthermore, O-698 demonstrated exceptional stability in cells, maintaining fluorescence for over 8 h, and was effective in monitoring RNA dynamics in response to RNA polymerase inhibition. Upon binding with RNA, O-698 offers a high signal-to-noise ratio for precise visualization without the need for wash steps. In 3D multicellular tumor spheroids, O-698 demonstrated robust penetration and uniform distribution throughout the tumor-like mass, reflecting its excellent capability to diffuse in dense biological tissues. In vivo imaging in tumor-bearing mice revealed strong fluorescence signals at the tumor injection site, consistent with the deep imaging capability of O-698. These findings underscore the potential of O-698 as a powerful tool for studying RNA dynamics in both cellular and in vivo environments. Overall, the rapid, stable, and wash-free imaging capabilities of O-698 make it a promising probe for real-time monitoring of RNA behavior, providing valuable insights into drug mechanisms and therapeutic efficacy. © 2025 Elsevier B.V.