Gut-liver-on-a-chip enables mechanistic study and risk assessment of drug-induced liver injury and drug-drug interactions.

Current preclinical models face challenges in recapitulating organ-level interactions affecting drug safety, and there has been little investigation into drug toxicity and related DILI. We present a pump-less gut-liver-on-chip, enabling integrated analysis of drug exposure-toxicity relationships and inter-organ pharmacological interactions. The platform incorporates a polarized intestinal barrier with a quadruple-cell co-cultured liver spheroid. Through simulation and comparative evaluation of oral versus systemic drug administration, we demonstrated the critical role of the intestinal barrier in modulating drug exposure, corresponding toxic responses and first-pass effects. Temporal profiling revealed progressive hepatic injury mechanisms involving mitochondrial dysfunction and activation of the apoptotic pathway. Pharmacological inhibition of cytochrome P450 attenuated victim-induced oxidative stress without affecting hepatic drug exposure, confirming enzyme-related bioactivation as the toxicity mechanism. Furthermore, transporter-mediated drug-drug interactions were functionally replicated, with perpetrator compounds altering substrate pharmacokinetics through competitive efflux inhibition and modified intestinal disposition. The ability of the platform to monitor drug exposure-toxicity relationships and drug-drug interactions was validated using combinations of perpetrator and victim drugs. This integrated approach advances applications of organ-on-chips by establishing causal relationships between drug exposure and toxicity, resolving the progression of temporal toxicity, and modeling drug-drug interactions, which are critical factors in predicting clinical hepatotoxicity and complex pharmacokinetic interactions.