Enzyme-Activated Biosensor Assisted by Enzymatic Rolling Circle Amplification for Sensitive Detection of APE1 and Imaging in Vivo

Precise identification of tumors is crucial for early diagnosis and treatment of cancer. However, spatially specific and sensitive molecular imaging of tumors remains a challenge due to issues such as low biomarker content and extra-tumor signal leakage. Endogenous purine/pyrimidine endonuclease 1 (APE1) is a marker for tumor diagnosis by detecting activity change and translocation from the nucleus into the cytoplasm. In this work, we developed a biosensor (named DNA/ZIF-8@Protein NPs) based on biomineralized metal-organic framework nanoparticles (MOF NPs) for sensitive detection and imaging of APE1 with the assistance of the rolling circle amplification reaction (RCA). In vitro experiments showed that based on the sensitivity of APE1 imaging, this biosensor can detect APE1 in the range of 0.005 to 2 U/mL with a detection limit of 0.0005 U/mL under optimal conditions. Functional probes and phi29 were loaded with ZIF-8, ensuring their simultaneous delivery to living cells. The results of in vivo experiments show that these DNA/ZIF-8@Protein NPs enable precise tumor cell identification and ultrasensitive molecular in situ tumor imaging by monitoring intracellular APE1. In summary, this method based on enzyme-activated RCA reaction provides new opportunities for ultrasensitive tumor in situ molecular imaging of tumor markers.