A microRNA‐21‐Activatable Two‐Stage Signal Amplification Probe for Catalytic Photodynamic Therapy

Photodynamic therapy (PDT) is noninvasive but lacks tumor specificity, while microRNAs (miRNAs) are highly specific biomarkers yet too low in abundance (pM-nM) to directly trigger therapy. Here we report ECyTz, a microRNA-21(miR-21)-activatable theranostic probe that achieves two-stage signal amplification. In the first stage, one miR-21 molecule catalytically activates multiple ECyTz probes through tetrazine-mediated transfer (TMT) bioorthogonal chemistry (turnover number 5.6-41 at 10-100 nM), because the probe releases the active photosensitizer upon reaction while the intact miRNA remains to trigger further cycles. In the second stage, each activated photosensitizer generates abundant reactive oxygen species (ROS). ECyTz remains non-fluorescent and photodynamically inert until miR-21 triggers a proximity-dependent inverse electron-demand Diels-Alder reaction, restoring near-infrared (NIR) fluorescence (27-fold) and robust ROS generation. The probe exhibits strict sequence specificity, minimal dark toxicity, and potent antitumor efficacy in xenograft models (86.7% tumor growth inhibition with no observable systemic toxicity), with upregulation of the downstream tumor suppressor PTEN confirming on-target activation. This modular miRNA-actuated platform with tunable recognition sequences establishes a proof-of-concept for catalytic bio-orthogonal signal amplification, offering a programmable strategy for precision tumor-selective photodynamic therapy.