A Tetrazine-Caged Carbon-Dipyrromethene as a Bioorthogonally Activatable Fluorescent Probe

A water-soluble 1,2,4,5-tetrazine-substituted carbon-dipyrromethene (C–DIPY) was synthesized from the previously reported carbonyl pyrrole dimer through a two-step procedure. Owing to the presence of a tetrazine moiety, the fluorescence emission of this compound was largely quenched in phosphate-buffered saline at pH 7.4. Upon addition of a bicyclo[6.1.0]non-4-yne (BCN) derivative, the tetrazine-based quenching component of the compound was disrupted through the inverse electron-demand Diels-Alder reaction to restore the fluorescence in up to 6.6-fold. This bioorthogonal activation was also demonstrated using U-87 MG human glioblastoma cells, in which the fluorescence intensity of this C–DIPY could be enhanced by 8.7-fold upon post-incubation with the BCN derivative. The results showed that this tetrazine-caged C–DIPY can serve as a bioorthogonally activatable fluorescent probe for bioimaging. The compound, however, was found to reside preferentially in the lysosomes instead of the mitochondria of the cells as predicted based on its cationic character, which could be attributed to its energy-dependent endocytic cellular uptake pathway, for which lysosomes are the end station. © 2023 Wiley-VCH GmbH.