Computational toxicology analysis of candidate drivers and pathways in HRZE-associated drug-induced liver injury.

The first-line anti-tuberculosis regimen HRZE (isoniazid (INH), rifampicin (RIF), pyrazinamide (PZA), ethambutol (EMB)) frequently causes drug-induced liver injury (DILI). However, the mechanisms underlying this combination hepatotoxicity remain unclear. Therefore, this study used a hierarchical network toxicology and molecular docking approach to systematically investigate the hepatotoxic mechanisms of the HRZE regimen, aiming to identify core and non-core hepatotoxic drugs and elucidate their synergistic effects. We identified 315 DILI-related targets associated with the HRZE regimen. Protein-protein interaction network analysis revealed seven hub genes, and enrichment analysis identified the biological processes involved and signaling pathways. Isoniazid, RIF, and PZA were classified as core hepatotoxic drugs due to the significant enrichment of their targets in DILI pathways and their high clinical risk. In contrast, EMB was identified as a non-core hepatotoxic drug, with lower target enrichment and a lower risk of liver injury. The targets of the core hepatotoxic drugs were widely distributed across major DILI pathways, while EMB may synergistically modulate pathway activity through a limited set of targets, including carbonic anhydrase II (CA2). Molecular docking confirmed that all four drugs bind stably to cytochrome P450 family 2 subfamily E member 1 (CYP2E1); the high affinity of EMB for CA2 suggests its involvement in cholestatic injury. In conclusion, HRZE-induced hepatotoxicity is a multi-pathway process driven predominantly by INH, RIF, and PZA, with EMB playing a synergistic role and potentially contributing to cholestatic injury. By establishing a multi-level toxicological framework, this study provides a theoretical basis for understanding combination hepatotoxicity and offers potential targets for DILI intervention.