Programmable DNA Circuit-Facilitated Determination of Circulating Extracellular Vesicle PD-L1 for Lung Cancer Diagnosis and Immunotherapy Response Prediction

Circulating extracellular vesicle (EV) PD-L1 is correlated with the occurrence and progression of lung cancer and has great potential as a valuable diagnostic and immunotherapy predictive biomarker. In this work, we propose a fluorescent biosensing method for the sensitive and accurate determination of circulating EV PD-L1. Specifically, after the phosphatidylserine-targeting peptide-assisted magnetic enrichment, a programmable DNA circuit is designed to translate the presence of PD-L1 to the appearance of numerous duplex DNA probes on the circulating EV surface. Upon fructose treatment, these newly formed duplex DNA probes are released from the EV surface to activate the trans-cleavage activity of CRISPR/Cas12a system, which finally produces a significant fluorescence signal. Experimental results reveal that the method not only enables sensitive determination of EV PD-L1 with a detection limit of 67 particles/mL but also demonstrates the potential use in the diagnosis and immunotherapy response prediction of lung cancer in a principle-of-proof study. Therefore, the method may provide a useful tool for EV PD-L1 determination, which may provide valuable information for the precise diagnosis and personalized treatment of lung cancer patients.