NIR FluorescenceStrategy for the Early Diagnosisof Melanoma Liver Metastasis Based on the Cascade Reaction of TwoSpecific Bioenzymes

Melanoma liver metastasis poses a serious risk to patient health, leading to deterioration of liver function accompanied by symptoms such as jaundice, abdominal pain, and weight loss. Early detection is paramount as it allows for timely intervention, potentially improving treatment efficacy and patient outcomes. Herein, a dual-target probe activated by butyrylcholinesterase (BChE) and tyrosinase (TYR) was proposed by initiating an enzyme cascade reaction, revealing NIR-emissive methylene blue (MB) as a responsive product. The cascade reaction mechanism was confirmed by enzyme activity inhibition experiments and high-resolution mass spectrometry. Intercellular imaging was carried out on the coincubation model of Hep G2 and B16 cell lines, which is the first attempt to the best of our knowledge. The results showed that BChE expressed in liver cells and TYR overexpressed in B16 cells jointly activated NIR fluorescence, which provides the possibility for the precise detection of melanoma liver metastasis. <i>In vivo</i> studies further proved that this method holds promise for the early diagnosis and postoperative adjunctive therapy of hepatic melanoma, ultimately enhancing survival rates.