Fluorescence Imaging-Incorporated Transcriptome Study of Glutathione Depletion-Enhanced Ferroptosis Therapy via Targeting Gold Nanoclusters.

Ferroptosis plays an important role in tumor inhibition and is a new type of programmed cell death. Recent studies have shown that glutathione (GSH) depletion is an effective method to enhance the therapeutic efficacy of ferroptosis; however, a systematic investigation of the phenomenon is limited. Herein, we provide a facile fluorescence imaging-incorporated transcriptome strategy to visualize the process and explore the mechanism of GSH depletion-enhanced ferroptosis. The proposed multifunctional nanoplatform is achieved using simple transferrin receptor aptamer-functionalized fluorescent gold nanoclusters (termed TfRA-AuNCs), which exhibit efficient hydroxyl radical generation and GSH-depleting capabilities. Live cell fluorescence imaging results revealed that TfRA-AuNCs were endocytosed into 4T1 cells and were mostly distributed in lysosomes. In vitro results indicated that TfRA-AuNCs enhanced the ferroptosis effect in 4T1 cells. Importantly, transcriptome analysis indicated that 4T1 cells treated with TfRA-AuNCs regulated the expression change of ferroptosis-related genes, and the Kyoto Encyclopedia of Genes and Genomes pathway identified the GSH metabolism pathway involved in ferroptosis, thus revealing the exact molecular mechanism of ferroptosis induced by TfRA-AuNCs at the RNA level. Furthermore, in vivo results confirmed the tumor inhibition effect, tumor-targeted fluorescence imaging, and long-term biocompatibility after TfRA-AuNC treatment. This study introduces a new possibility for the mechanistic study of nanoagent-induced ferroptosis in tumor treatment.