a) Gold-triggered activation of prodrugs 4 a–c in A549 cancer cell culture. Negative controls: [Au]-resins (1 mg mL⁻¹); 4 a–c (10, 100, and 1 μm, respectively). Positive control: 5 a–c (10, 100, and 1 μm, respectively). Prodrug activation assay: [Au]-resins+4 a–c (10, 100, and 1 μm, respectively). Cell viability was measured at day 4 using PrestoBlue reagent. Error bars: ±SD from n=3. b) Bioorthogonal gold-mediated release of green fluorescent Rhodamine 110 from precursor 1 in the brain of a zebrafish. The presence of the [Au]-resin is indicated with a white arrow. Study of fluorescence intensity shows high statistical significance compared to the negative control (DMSO).

Time-course analysis and representative images of the bioorthogonal gold-mediated release of green fluorescent rhodamine 110 from precursor 1 in the brain of a zebrafish. Study of fluorescence intensity between the persistent (group A, in black) vs the intermittent (group B, in grey) treatment at 3 dpt shows statistical significance. This study further corroborated previous observations in cancer cell culture. Persistent treatment with reagent 1 (group A), enabled the local generation of dye 2 in the brain and the sustenance of high levels of fluorescence over the life of the study. In contrast, after interrupting the treatment for 2 d (group B), local levels of fluorescence decreased, likely due to the metabolization / elimination of compound 2. 24 h after resuming the treatment (3 to 4 dpt), high intensity of local fluorescence in the area of the [Au]-resin was restored. Importantly, treatment with reagent 1 in the absence of [Au]- resin did not change fluorescence levels in the brain over the background control.