SPECTRA-based Detection of Drug-Induced Hepatotoxicity through Extracellular Vesicle Analysis.

Extracellular vesicles (EV) are becoming crucial targets in liquid biopsy, diagnostics, and therapeutic applications, yet their nanoscale characterization remains challenging. In this context, the detection of drug-induced liver injury, i.e., hepatotoxicity, through EV molecular content remains unexplored. To this end, we present the SPECTRA - Surface Plasmonic Enhancement with Combined Transformative RAman and Interferometric Microscopy approach, which provides rapid, label-free EV content analysis under 30 minutes and requires only 1.3 microliters of sample. Using hepatic cultures as a model, our platform captures distinct and reproducible EV molecular changes in response to acetaminophen-induced hepatotoxicity. Across independent culture sets, a Gaussian Process Regression model predicted Acetaminophen (APAP) dose with median absolute error of 1.50 mM and minimal bias (Bland-Altman bias ~0.03 mM); cross-validated RMSE was 3.145 mM. These findings establish EVs as dynamic reporters of cellular drug responses and demonstrate use of SPECTRA for EV detection of hepatotoxicity.